Cultured plant cell gums for food, pharmaceutical, cosmetic and industrial applications

ABSTRACT

Certain cultured plant cell gums, including those produced in suspension culture of plant cells of plants of the family Aizoaceae are described. Plant cell gums of plants of the genus Mesembryanthemum are specifically provided. Also described are the methods of using these cultured plant cell gums in the manufacture of food products, pharmaceuticals and veterinary products, cosmetics and other industrial products, such as paper, adhesive, ink, textiles, paint, ceramics, explosives, cleaning agents or detergents, products for firefighting, agricultural chemicals including pesticides and fungicides, for oil and gas production, and in photography, lithography, and other industries are described. Food, pharmaceutical, veterinary, industrial and cosmetic compositions containing certain cultured plant cell gums are also described. Plant cell gums can be employed as substitutes for plant exudate and extract gums and other known emulsifying, viscosifying and gelling agents.

RELATEDNESS OF THE APPLICATION

This application is a continuation of U.S. patent application Ser. No.09/072,568, filed May 5, 1998, now U.S. Pat. No. 6,271,001 which in turnis a continuation of U.S. patent application Ser. No. 08/409,737, filedMar. 23, 1995, now U.S. Pat. No. 5,747,297, which are incorporated intheir entirety by reference herein.

FIELD OF THE INVENTION

The subject invention relates generally to the use of cultured plantcell gums in food, pharmaceutical, cosmetic and other industrialapplications, including their use in oil and gas well drilling andproduction and lithography, and in the manufacture of textiles, ink,adhesive, paper, paint, ceramics, agricultural chemical and cleaning ordetergent agent

BACKGROUND OF THE INVENTION

A variety of natural and semisynthetic complex carbohydrates orpolysaccharides have been commercially important in human and pet foodmanufacturing; in the cosmetic, paper, textile, paint, agricultural,explosives, hydrolube, adhesive, ceramic, cleaning polish, detergent,fire fighting, ink, photography, lithography, and deodorant gelindustries; and in mining, and gas well drilling and production. Naturalcomplex carbohydrates and polysaccharides include seaweed extracts,plant exudates, seed or root extracts, and microbial polysaccharidesproduced by fermentation. Semisynthetic complex carbohydrates andpolysaccharides include cellulose derivatives, low-methoxyl pectin,propylene glycol alginate, triethanolamine alginate and guar gumderivatives. Sandford, P. & Baird, J. (1983) “Industrial Utilization ofPolysaccharides” in The Polysaccharides, Vol. 2, pp. 411-491.

The production of natural complex carbohydrates or polysaccharides isfrequently problematic. For plant exudates and seed or root extracts,production is dependent on climate and harvest conditions. For example,gum arabic is an exudate from Acacia senegal trees. Gum production isstimulated by stripping the bark from the trees; the gum is collected byhand in the form of “dried tears.” Production of gum arabic can varyeach year as a function of weather conditions, labor strikes, naturaldisasters, etc. Meer et al. (1975) Food Technology 29:22-30. Theunreliable supply results in variable gum arabic cost. Seed gums, suchas guar gums are expensive due to harvesting costs. Guar gum is derivedfrom the seed of the guar plant Cyamopsis tetragonolobus. Processinginvolves removal of the seed coat, separation of the germ from theendosperm, and milling of the endosperm. Sandford, P. & Baird, J.(1983), supra. Further, gums obtained from such sources may havevariable quality and exhibit variable functional properties.

The production of seaweed extracts can also be problematic. Agarproduction is labor intensive in that it involves the harvesting of redseaweed by hand: in some areas of the world, divers in full pressuresuits collect individual plants in deep water; in other places, theseaweed can be collected at low tide without the use of divingequipment. Carrageenan or Irish Moss is produced from another redseaweed harvested by raking and hand gathering. Algin is produced frombrown algae which can be harvested manually or with small mechanicalharvesters. Sandford, P. & Baird, J. (1983), supra.

Further, hand harvesting can introduce a purity problem. For example,hand collected lots of gum arabic are seldom pure; samples areclassified according to grade which depends on color, and contaminationwith foreign bodies such as wood or bark (VanNostrand's ScientificEncyclopedia, 7th ed. (1989) D. Considine (ed.), Vol. I, p. 1389).

Microbial fermentation gums such as xanthan gum avoid many of thedifficulties associated with harvesting of plant exudates or extractionof algae because production is carried out in fermentation facilities.However, xanthan gum production poses other problems. Xanthan gum isproduced by Xanthamonas campestris, which presents a cell disposalproblem because X. campestris is a plant pathogen (Scaad, N. W. (1982)Plant Disease 66(10):882-890). Xanthan gum has also been objected to asbeing too expensive for certain applications such as drilling mud. See,e.g., Kirk-Othmer Chemical Engineering Encyclopedia (3rd. ed. 1981)17:153.

Thus, there is a clear need in a number of industries for a reliable,relatively inexpensive gum or class of gums that do not create adisposal problem. While a number of plant cells have been observed toproduce polysaccharide and/or complex carbohydrates when cultured(Aspinall, G. & Molloy, J. (1969) Canadian J. Biochem. 47:1063-1070;Fincher, G. et al. (1983) Ann. Rev. Plant Physiol. 34:47-70; Clarke, A.et al. (1979) 18:521-540; McNeil, M. et al. (1984) Ann. Rev. Biochem.53:625-663; Hale, A. et al. (1987) Plant Cell Reports 6:435-438; andBacic, A. et al. (1987) Australian J. Plant Physiol. 14:633-641), it hasnot been suggested that such cultured plant cell gums might be suitablein the pharmaceutical, paper, textile, paint, agricultural, explosives,hydrolube, adhesive, ceramic, cleaning polish, detergent, fire fighting,ink, photography and lithography industries; or in mining, and oil andgas well drilling and production. Only Otsuji, K. et al. EP 0 285 829(published Oct. 12, 1988) have utilized cultured Polianthus gum incosmetic applications.

Related work by the inventors hereof has been published in WO 8806627(1988) and WO 9402113 (1994). WO 8806627 relates in general to the useof cultured plant cell gums in the manufacture of food products asemulsifiers, thickening agents, gelling agents and the like. Culturedplant cell gums of Pyrus, Prunus, and Rosa are specifically exemplified.U.S. Pat. No. 5,133,979 (issued Jul. 28, 1992) and U.S. Pat. No.5,296,245 (issued Mar. 22, 1994) are directed to similar subject matter.WO 9402113 relates to the general use of cultured plant cell gums asemulsifiers, viscosifiers, and the like for the manufacture ofindustrial, pharmaceutical or cosmetic products. Cultured plant cellgums from suspension cultures of Nicotiana, Pyrus, Phleum and Lolium areexemplified

SUMMARY OF THE INVENTION

The subject invention comprises the use of cultured plant cell gumsproduced from gum-secreting cells of vascular plants in a variety offood, pharmaceutical, veterinary, cosmetic and industrial applicationsincluding, without limitation, textiles, paper, adhesives, inks,lithography, ceramics, cleaning detergents, firefighting, agricultural,explosives and oil and gas wells. The cultured plant cell gums areuseful in general as viscosifiers, as thickening, gelling, emulsifying,dispersing, suspending, stabilizing, encapsulating, flocculating,film-forming, sizing, adhesive, texture-modifying, enrobing, bindingand/or coating agents, and/or as lubricants, water retention agents andcoagulants. Any cultured plant cell gum can be useful in the subjectindustrial, pharmaceutical and cosmetic applications described herein.This invention is more specifically directed to particular plant cellgums which have particularly useful rheological properties and the useof such gums in food, pharmaceutical, veterinary, cosmetic and otherindustrial applications. The invention also relates to several specificapplications for which cultured plant cell gums are particularlysuitable.

In one embodiment the invention relates to cultured plant cell gumsecreted in culture of plants of the family Aizoaceae. This family ofsucculents includes plants of the genera: Mesembryanthemum, Aptenia,Carpobrotus, Delosperma, Hereroa and Rushia, among others. Of moreinterest are plant cell gums of the Mesembryanthemum, Aptenia, orCarpobrotus. The plant cell gums of this family are particularly usefulas emulsification agents and emulsion stabilizing agents. The plant cellgums of these plants are very active emulsifiers. Plant cell gums ofspecies of Mesembryanthemum are particularly useful in the formation oflow viscosity, low-droplet-size emulsions, e.g., cloud emulsions.Low-droplet-size emulsions find extensive use, for example in the foodindustry, for manufacture of soft drinks. Methods of use of thesecultured plant cell gums are provided.

In another embodiment the invention relates to cultured plant cell gumsecreted in culture of monocot plants, including plants of the familyPoaceae including plants of the genera Phleum and Panicum, among others.Cultured plant cell gums of Phleum (particularly those of timothy grass,P. pratense) exhibit good gelling ability and high viscosity. Phleumcultured plant cell gum can serve as a substitute for guar gum orhydroxymethylcellulose. Panicum gums exhibit high viscosity andvisco-elastic properties and have a variety of applications in the foodand other industries, particularly for the preparation of drilling muds.Panicum cultured plant cell gums are useful in the manufacture ofchemical sprays, particularly for agricultural sprays to inhibitsatellite droplet formation in such sprays. Methods of use of thesecultured plant cell gums are provided.

Table 1A provides a preferred list of families, genera and species ofplants that are useful in for the production of cultured plant cellgums. Table 1B provides a list of more preferred families, genera andspecies of plants useful for production of cultured plant cell gums.Plant families of more interest for production of cultured plant cellgums include: Actinidaceae, Agavaceae, Aizoaceae, Asteraceae,Cucurbitaceae, Fabaceae, Malvaceae, Mimosaceae, Poaceae, Rosaceae, andSolanaceae. Plant genera of more interest for production of culturedplant cell gums include: Acacia, Actinidia, Carpobrotus, Chichorium,Cucumis, Glycine, Hibiscus, Hordeum, Letuca, Lycopersicon, Malus,Medicago, Mesembryanthemum, Nicotiana, Oryza, Panicum, Phalaris, Phleum,Polianthus, Pyrus, Rosa, Sida, Solanum, Trifolium, Trigonella, and Zea.Plant species of more interest for production of cultured plant cellgums include: Acacia senegal, Actinidia deliciosa, Carpobrotus spp.,Chichorium intybus, Cucumis sativus, Glycine max, domesticus, Medicagosativa, Mesembryanthemum spp., Oryza sativa, Panicum miliaceun, Hibiscusesculentus, Hordeum vulgare, Letuca sativa, Lycopersicon esculentum,Malus domesticus, Phalaris aquaticus, Phleum pratense, Polianthustuberosa, Rosa glauca, Sida rhombifolia, Solanum, Trifolium repens,Trifolium pratense, Trigonellafoenum-graceum, and Zea mays. Culturedplant cell gums produced by gum-secreting cells of plants of theforegoing families, genera and species are useful as emulsifying agents,viscosifying agents, gelling agents, thickening agents, dispersing orsuspending agents, emulsion stabilizing agents, encapsulating agents,flocculating agents, film-forming agents, sizing agents, binding and/orcoating agents, and/or as lubricants, water retention agents andcoagulants or in adhesive compositions and the like in food,pharmaceutical, veterinary, cosmetic and other industrial applications.Methods of use of these cultured plant cell gums are provided.

In general, plant cell lines that produce at least about 0.05% (w/v) gumin the final fermentor culture broth, are preferred to reduce productioncosts. Plant cell lines that produce at least about 0.5%, 2.0%, and10.0% (w/v) gum in the final culture broth are increasingly preferred.In one embodiment, the cultured plant cell gums employed in suchapplications are cultured plant cell gums having arabinogalactanproteins (AGPs) of at least about 4.0% (w/w). As discussed herein,choice of explant and culture conditions for the plant cells can affectfunctional properties of the gum product.

Cultured plant cell gum products can be used as a substitute for priorart gums, such as gum arabic and guar gum. The cultured plant cell gumscan also be used as a substitute for xanthan gum, alginic acid, agar,calcium alginate, carrageenan, guar gum, karaya gum, locust bean gum,potassium or sodium alginate, tragacanth gum and others. For example,the cultured plant cell gums can be used as thickening agents and/oremulsifying agents to replace gum arabic in adhesives, inks, textileprinting and cosmetics. The cultured plant cell gums can be used toreplace alginic acid as an emulsifier, thickening agent, suspendingagent, waterproofing agent, etc. in toothpaste, cosmetics,pharmaceuticals, textile sizing, coatings, oil-well drilling muds, andconcrete. The cultured plant cell gums can be used to replace agar as agelling agent, protective colloid, in photographic emulsions or otherapplications. The cultured plant cell gums can be used to replacecalcium alginate as a thickening agent, stabilizer, etc. in syntheticfibers. Carrageenan, which can be used as an emulsifier, protectivecolloid, stabilizing agent, etc. in toothpastes, cosmetics andpharmaceuticals, can be replaced by cultured plant cell gums. Culturedplant cell gums can substitute for guar gum, which functions as athickening agent, emulsifier, etc. in paper, cosmetics, pharmaceuticals,textiles, printing, polishing, and as a fracture aid in oil wells.Cultured plant cell gums can also replace karaya gum as a protectivecolloid, stabilizer, thickener, emulsifier, etc. in pharmaceuticals,textile coatings and adhesives. Cultured plant cell gums can replacelocust bean gum (carob-bean gum) as a stabilizer, thickener, emulsifier,etc. in packaging material, cosmetics, sizing and finishes for textiles,pharmaceuticals and paints. Potassium or sodium alginate, which canfunction as an emulsifier, thickening agent, stabilizer, etc. inpharmaceuticals, textile printing, cement compositions, paper coatings,and in some water-base paints, can be replaced by cultured plant cellgums. Cultured plant cell gums can replace tragacanth gum as anemulsifying agent, coating agent, thickening agent, stabilizer, etc. inpharmaceuticals, adhesives, leather dressings, textile printing andsizing, dyes, toothpastes, hairwave preparations, soap chips andpowders. Xanthan gum, which is used as a thickening, suspending,emulsifying agent, stabilizing agent, etc. in oil and gas well drillingmuds and other applications, can also be replaced by cultured plant cellgums. In replacing such prior art gums, the cultured plant cell gums canoffer unexpectedly improved results. Often, cultured plant cell gums cansurprisingly be used in smaller quantities than the prior art gums toachieve equivalent functional results. Further, production of thecultured plant cell gums do not present the cell disposal problem thatxanthan gum production does.

The cultured plant cell gums are not useful in applications where theirutilities or properties are significantly compromised or destroyed.Organic solvents such as alcohol, acetone and ether and the like candisrupt function by causing precipitation of the cultured plant cellgums. To maintain the gums' emulsification, thickening or gellingproperties, it is preferred that the temperature of the gum-containingsolution or mixture be maintained between about 4° and 90° C. and have apH of neutral to slightly alkaline. As the pH increases, the thickeningcapacity of the gums decreases. However, even at elevated pH, viscositycan increase with increased ionic strength. Gum-containing solutions cangel in the presence of divalent cations such as calcium, and astemperature decreases, gel strength increases. Typically, stable gelsare produced in the pH range of between about 3 to 10 and in thepresence of calcium ions. Further, heating and cooling of gelled gumsolutions between ambient and 80° C. has not reduced gel strength,indicating that the gels can be thermo-reversible.

The cultured plant cell gums of this invention are useful in a widevariety of applications in part because they are stable over a widerange of temperatures. In an emulsion or solution, the gums arefunctional over a temperature range of about 0° to 100° C. at neutralpH. The dried gum powder (neutral pH) is stable over a temperature rangeof about −70° C. to about 10° C. If heated, the dried, powdered gum cancaramelize. Furthermore, cultured plant cell gums of this invention canprovide substantially non-toxic rheological agents (emulsifiers, etc.)of biological origin that can replace potentially harmful syntheticpolymers and surface active agents.

The invention also provides isolated (i.e., substantially free of cellbiomass) cultured plant cell gums which may be provided as aqueoussolutions or suspensions (more or less concentrated than in culturefiltrate) or as dried powders. Isolated plant cell gums of thisinvention include those of plants of the genera: Acacia, Actinidia,Aptenia, Carpobrotus, Chickorium, Cucumis, Glycine, Hibiscus, Hordeum,Letuca, Lycopersicon, Malus, Medicago, Mesembryanthemum, Oryza, Panicum,Phalaris, Phleum, Polianthus, Sida, Solanum, Trifolium, Trigonella, andZea. Of particular interest are isolated cultured plant cell gums ofplants of the family Aizoaceae (including those of the generaMesembryanthemum, Aptenia, and Carpobrotus).

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph illustrating the relative emulsification capacity ofthe Mesembryanthenum plant cell gum (squares) compared to a commercialsprayed-dried gum arabic (circles). Droplet size in )m is plotted on alog scale as a function of gum concentration (wt %).

DETAILED DESCRIPTION OF THE INVENTION

The present work is an extension of the work disclosed in WO 8806627(1988) and WO 9402113 (1994) which disclosed the general ability ofcultured plant cell gums to function as emulsifying agents, thickeners,stabilizers, texture modifiers, gelling agents, binding or coatingagents, suspending agents or the like. The present work specificallydescribes and exemplifies additional sources of and specializedapplications of certain cultured plant cell gums.

The invention also provides isolated (i.e., substantially free of cellbiomass) cultured plant cell gums which may be provided as aqueoussolutions or suspensions (more or less concentrated than in culturefiltrate) or as dried powders. As will be appreciated in the art,isolated plant cell gums for use in food and veterinary applicationsshould be substantially free of harmful levels of toxic plant components(oxalates, alkaloids and the like).

“Cultured plant cell gum” is defined as the substantially cell-freematerial recovered from cultured plant cells, and is usedinterchangeably herein with “gum product.” The cultured plant cells arethose which are capable of synthesizing components of the gum productand transporting the same extracellularly in culture. A variety ofvascular plant cells, including those derived from gymnosperms andangiosperms, may be used in the subject method (see Table 1A and 1B).Cells of plants of the Dicotyledonae class (e.g., the Rosidae andAsteridae subclasses) and Monocotyledonae class (e.g., the Commelinidaesubclass) can be used in the subject methods. Table 1A provides a listof preferred families, genera and species of plants useful in thepreparation of cultured plant cell gums. Table 1B provides a list ofmore preferred families, genera and species of plants useful in thepreparation of cultured plant cell gums.

The cultured plant cell gum comprises complex carbohydrates andoptionally glycoproteins, which are secreted into the medium by thecultured cells. The major classes of complex carbohydrate polymers areproteoglycans (e.g., arabinogalactan proteins (AGPs)), polysaccharides(e.g., neutral and acidic pectins), hetero- and homo-glucans,heteroxylans, and hetero- and homo-mannans (McNeil et al. (1984) Ann.Rev. Biochem. 53:625-633). Complex carbohydrates and glycoproteins areknown to be secreted by many cultured cell lines (Clarke, A. et al.(1979) Phytochemistry 18:521-540; Fincher et al. (1983) Ann. Rev. PlantPhysiol. 34:47-70; Bacic, A. et al. (1987) Australian J. Plant Physiol.14:633-641). Monocot gum-secreting plant cells have been found tosecrete gums containing a root-slime-like material which contributes tothe functionality of the gum as providing good gelling capacity and/orenhanced viscosity and/or visco-elastic properties compared in generalto dicot gums.

The cells to be cultured can be initiated from a variety of explants,for example, a leaf, style, anther or stem of a plant, segments of whichcan be placed on solid plant culture medium. Callus cells mayproliferate from any of the tissues of these organs and the callus cellscan then be transferred to liquid suspension culture. Alternatively,seeds can be surface sterilized, and placed in a solid or liquid planttissue culture medium to initiate germination. The germinating seedlingscan then be maintained, for a time, in liquid suspension culture. Thesuspension culture medium can be any known suitable medium such as MSmedium (Mirashige, T. & Skoog, F. (1962) Physiologia Plantarum15:473-497; Wu, M. & Wallner, S. (1983) Plant Physiol. 72:817-820). orvariants thereof. Suspension culture medium can be optimized forenhanced cell growth and/or enhanced gum production. Transfer tosuspension culture is preferred because in general it increases gumproduction and because it is possible to scale up a liquid suspensionculture. Air fermentors are preferred because they reduce shear stresson the cells. While cells can produce gum on a solid medium, massculture on solid media poses a number of practical difficulties,including gum collection.

Usually, a plant cell hormone is employed to enhance cell growth and/orpolysaccharide production. Plant hormones include, for example, theauxins such as 2,4-dichlorophenoxyacetic acid (2,4-D), naphthoxyaceticacid (NOA) and 2,4-dichlorophenoxybutyric acid (2,4-DB) or mixturesthereof. The use of given hormone may provide improved callus or cellgrowth for a given plant cell. For example, NOA gave improved resultsover 2,4-D for callus and cell cultures of Medicago sativa. Plant celllines can be adapted using methods well-known in the art to exhibit goodgrowth on lower levels of (or no) plant hormones. Using such adaptationmethods, callus and suspension cultures of Pyrus and Mesembryanthemumthat grow well on media containing no 2,4-D have been obtained. The useof lower hormone levels in suspension culture can decrease gummanufacture cost and alleviate the potentially detrimental effects ofhormones on plant cells.

The methods described herein for generation of callus and the initiationand maintenance of suspension cell cultures are generally applicable orreadily adaptable, employing art-known media, methods and expedients, tocells of any vascular plant including those listed in Table 1A and 1B.The specific culture conditions for Nicotiana. plumbaginifolia, Pyruscommunis and Phelum pratense are exemplified herein.

A variety of growth media suitable for the various plant cells of thisinvention are known in the art. A variety of carbon sources can bereadily employed (or cells can be readily adapted for growth on a givencarbon source) including sucrose, glucose, fructose and lactose. Morecomplex carbon sources can be employed, e.g. double enzyme hydrolyzedglucose syrup or brewers' liquid maltose (BLM). Carbon source may affectthe Theological properties of cultured plant cell gums. It has beenobserved that employing BLM as a carbon source increases cell growth andgum yield for certain types of cells. Growth of Nicotiana on BLM resultin production of a plant cell gum having excellent film-formingproperties. It has also been observed that an increase in osmoticpressure or in sucrose concentration in the medium can increase gumproduction by some cultured plant cells.

Addition of other additives to growth media may affect the viscosity orother properties of plant cell gums. For example, growth of a Nicotianasuspension on 5% glycerol, believed to function as an osmodicant in themedium, resulted in cultures having significantly higher viscosity.

The gum product can be recovered from the culture medium by methods wellknown in the art. See Johns, M. & Noor, E. (1991) Aust. J. Biotechnol.5(2):73-77; Golueke, C. et al. (1965) U.S. Pat. No. 3,195,271; Seviour,R. & Kristiansen, B. (1983) Eur. J. Appl. Microbiol. Biotechnol.17:178-181; Mort, A. et al. (1991) Carbohydrate Res. 215:219-237; andWu, M. & Wallner, S. (1983) Plant Physiol. 72:817-820. A specificrecovery and purification method is exemplified herein. A “complexant”is a composition or compound that sequesters calcium or other divalentmetal ions from the gum product during the recovery procedure. Forexample, Na₂. EDTA added during the recovery process chelates calcium.Other sequestering agents such as citrate, cyclohexane diaminetetraacetate (CDTA), imidazole, sodium hexametaphosphate may also beused. Sequestering of calcium is desirable to avoid the formation ofinsoluble complexes during drying of the recovered gum. Preservativesand antioxidants can be added to the culture filtrate to minimize gumdegradation.

As noted above, plant cell gums are complex mixtures includingcomponents having a range of molecular weights. Molecular weightfractionation methods can be applied during plant cell gum isolation orto isolated plant cell gum to obtain gum fractions of differentmolecular weight range. These fractions can have distinct Theologicalproperties, e.g., lower molecular weight fraction may exhibit generallylower viscosity.

In one embodiment this invention relates to cultured plant cell gums ofplants of the family Aizoaceae including plants of the generaMesembryanthemum, Aptenia and Carpobrotus. These genera are closelyrelated and species once classified in one of these genera may currentlybe classified in another of these genera, for example Mesembryanthemumchilense has been reclassified as Carpobrotus chilense. This inventionincludes plant cell gums of all plants classified into these genera,including varieties of each and plants that may result from a crossbetween two species of the genera. More specifically this inventionincludes cultured plant cell gums (as well as uses thereof) of plantsdesignated as Mesembryanthemum spp., including M. crystallinum, M.edulis, M. nodiflorum, M. barklyi, M. criniflorum, M. forsskalei Hochst,M. cordifolium and other species listed in Table 1B, Carpobrotus spp.,including Carpobrotus chilense, Carpobrotus acinaciformis, Carpobrotusedulis, Carpobrotus aequilaterus, Carpobrotus modestus, Carpobrotusmuirii and other species listed in Table 1B, Apentia cordifolia, Apteniacordii, varieties of each and plants resulting from crosses between twospecies. See: H. Jacobsen (1974) Lexicon of Succulent Plants BlandfordPress Ltd. (London) and H. Jacobsen (1960) Handbook of Succulent PlantsBlandford Press Ltd. (London) for descriptions of Aizoaceae includingadditional species of Mesembryanthemum, Aptenia and Carpobrotus.

The skilled practitioner, using information available in the art and theteachings of the subject application, can identify cultured plant cellgums that are useful as thickening, gelling, emulsifying, dispersing,suspending, stabilizing, encapsulating, flocculating, film forming,sizing, adhesive, binding and coating agents, and as lubricants, waterretention agents and coagulants, etc. in the aforementioned industries.The suitability of using a cultured plant cell gum for a particularapplication can be assessed by methods known to those of skill in theart.

The cultured plant cell gums can be used to establish and stabilizesolid, liquid and gaseous dispersions. An emulsion is an intimatemixture of two invincible liquids in which one phase is dispersedthroughout the other as small, discrete droplets (Sandford, P. & Baird,J., “Industrial Utilization of Polysaccharides” in The Polysaccharides(1983), Academic Press, Inc., Vol 2, pp. 411-491). The cultured plantcell gums can be used as emulsifying agents or stabilizing agents inemulsions. Suspensions are solid particles dispersed uniformlythroughout a liquid phase (a suspension) mainly by increasing theviscosity of the suspension liquid phase with suspending agent. Foamsare gas dispersed in a liquid or solid phase. When cultured plant cellgums are employed as foam stabilizers, they affect the surfaceproperties (e.g., interfacial tension) of foams, thereby promoting afirm, stable foam.

Emulsification capacity can be assessed by, for example, measuring thereduction in aqueous surface tension or interfacial tension due to thegum product, measuring the critical micellar concentration (CMC), ormeasuring the hydrophile-lipophile balance (HLB; the ratio of polar tononpolar portions of the composition). Additional methods of assessingemulsifying capacity include particle sizing and counting, and effect onviscosity and electrical properties of the emulsion due to the gumproduct. For a discussion of such methods, see Zajic, J. & Panchal, C.in CRC Critical Review in Microbiology (1976), pp. 39-66. The choice ofa particular gum product for a desired application depends on additionalfactors such as solubility and compatibility with other chemicals in theemulsion mixture, and pH, ionic strength and temperature of the emulsionmixture. The specific method employed to measure the emulsificationcapacity for at least some of the gum products described herein involvesmeasurement of turbidity and droplet size and is described in theExamples.

Emulsion stabilizing capacity is the ability of a gum to maintain anemulsion over time. Emulsion stability can be tested by evaluating theturbidity of the emulsion (or industrial emulsion mixture) over time.

Thickening agents increase the viscosity of aqueous solutions orsuspensions. They increase the resistance to flow of a liquid. Sandford,P. A. & Baird, J., supra. Viscosity imparted by cultured plant cell gumsto mixtures or solutions can be measured with commercially availableviscometers. Such viscometers commonly employ methods based on Stoke'slaw, the capillary tube method, the rotating cylinder method or theoscillating disk method. The specific method employed to measure theviscosity of at least some of the gum products described herein isdescribed in the Examples.

Assessment of gelling capacity of a gum product can be carried out bymethods known in the art. The specific method employed to measuregelling capacity of at least some of the gum products described hereinis set forth in the Examples. Gelling capacity can be assessed bymeasuring the rupture strength, shear modulus, back extrusion andmelting and setting points of the gum product.

Lubricating capacity can be assessed by methods known in the art. Forexample, an adaptation of ASTM (American Society for Testing Materials)Method D4172 may be used.

Encapsulating capacity can be assessed by methods known in the art. Aspecific method is described in Example 3 of the U.S. Patent Applicationfor “Plant Gum Material and Use Thereof in Food Products,” filed on evendate herewith.

It is desirable in certain emulsification applications, particularly inthe manufacture of compositions of agricultural chemicals for sprayapplications, to inhibit satellite droplet formation and thereby tominimize undesired dispersion of potentially hazardous materials. Theability of a plant cell gum to provide such inhibition can be tested byassessing splash inhibition in solutions/compositions containing thegum. Splashing behavior of droplets can be assessed by video monitoringof drop impact as is known and understood in the art.

In some cases, particular functional properties have been associatedwith particular gum components. It has been observed that AGP in thecultured plant cell gum product can enhance emulsification properties.For example, Pyrus communis and Nicotiana plumbaginifolia have higherlevels (6-11% (w/w)) AGPs, while Phleum pratense produces a gum withnondetectable AGP and poor gelling and emulsification capacity. Phleumpretense has comparable viscosity to Pyrus and Nicotiana gums withoutthe gelling and emulsification properties. Phleum pratense is thususeful as a viscosity enhancer in applications where emulsification isnot desired, e.g., in applications where guar gum andhydroxymethylcellulose have traditionally been used.

Those embodiments of the subject invention which use the gum products asemulsifiers preferably employ a gum product relatively rich in AGPs. Inparticular, cultured plant cell gums containing at least about 4% (w/w)AGP in the gum can be useful. Complex carbohydrates in the culture fluidcan be determined by the method of Dubois et al. (1956) Anal. Chem.28:350-356. AGP can be determined by the method of Van Holst, G. &Clarke, A. (1985) Anal. Biochem. 148:446-450. AGP-containing gums havebeen found in higher plants (14 orders of angiosperms, 3 orders ofgymnosperms), and in lower plants (e.g., Fontinalis anti-pyretica).Fincher, G. et al., supra.

It has been found that a gum product recovered from Pyrus communis cellssuspension cultured in MS medium plus 2,4-D has complex carbohydrates atabout 5.26 mg/mL of culture fluid as determined by the method of Duboiset al. (1956) Anal. Chem. 28:350-356; and 8.9% (w/w) AGP as determinedby the method of Van Holst et al. (1985) Anal. Biochem. 148:446-450.

The cultured (MS medium) gum product of Lolium multiflorum and Nicotianaplumbaginifolia have been found to have an AGP % (w/w) of 11.0 and 4.5,respectively. In contrast, cultured cells (MS medium) of Phleum pratensehave been found to have no detectable AGP (detection limit is about 0.25μg by the method of Van Holst et al. (1985)).

A description of particular applications in which the cultured plantcell gums can be employed follows. This discussion is not intended to belimiting.

Applications of cultured plant cell gums for food applications have beendescribed in WO 8806627 and U.S. Pat. Nos. 5,133,979 and 5,296,245. Forexample, plant cell gums of this invention are useful in the preparationof oil-free salad dressing and as whipping bases for Gil mousses,desserts, yoghurts and fruit-based foams, as an encapsulating agent forflavors (e.g., flavors [garlic, onion, herbs, “smoky” flavors and“cheese” flavors] and flavor oils [orange and lemon oils]), film-formingagents for enrobing or coating foods, and thickening agents, for examplefor, sauces, toppings, spreads, fillings, dips, custards and gravies.

Gellan is currently employed as a film forming ingredient in bread crumbmixes and batters for coating various food items (e.g. meats, cheese,fish) or dough-enrobed foods (egg rolls) for frying to increasecrispness and decrease oil absorption or to give items that are heatedby microwave a “deep-fried” appearance. Cultured plant cell gums of thisinvention can be employed in such applications as a substitute forgellan. Film made from plant cell gums can exhibit excellent resistanceto the application of dry heat (up to about 100° C.) making them usefulin various cooking applications.

Certain plant cell gums (e.g., that of Pyrus) can exhibit significanttemperature stability such that gels prepared with the gum do not meltor soften up to temperatures of about 80° C. These gums are useful inthe preparation of non-melting sauces, e.g., barbecue, cheese or buttersauces and other sauces for cooking.

Cultures plant cell gums can exhibit rapid cold gelling in cold watermedia. This property is useful in the production of instant dehydratedsauces, toppings, spreads, fillings, dips, custards, gravies and thelike.

Gelatine is used extensively for preparing capsules for human andveterinary application, including drug, vitamin and food supplementapplications. Gelatine is also used for the production of oil-filledcapsules for nutrient oils including evening primrose oil, cod liveroil, vitamin E and the like. Films prepared from certain plant cellgums, including gums of Pyrus, are oil-resistant exhibiting little or nodeterioration on exposure to oil. Cultured plant cell gums can beemployed to prepare films that can be substituted for gelatine incapsule applications, particularly for use in oil-filled capsules.Oil-resistant gum film can also be employed to enrobe oily foods, e.g.,gum films can be employed to enrobe nuts for subsequent coating withchocolate.

Plant cell gums of this invention can be employed as dietary fibersupplements.

Plant cell gums are useful in a variety of veterinary applications forpreparation of food supplements and vitamins and in the preparation ofslow release pellets that can be used to deliver vitamins or drugs(anthelminthic). Plant cell gums, particularly those that exhibitheat-stable gel formation, can be employed as gelling agents in themanufacture of canned pet food.

In the paper industry, prior art gums have been used in wet end beateraids, surface sizes (e.g., size press and calender), pigmented coatings(e.g., blade, roll airknife, and size press coatings), and in adhesives.Sandford, P. & Baird, J., supra. Cultured plant cell gums can be used assubstitutes for such prior art gums as locust bean gum, karaya and guargums as hydrophilic colloids employed in the wet end as beater aids toreduce flocculation of pulp suspensions and improve paper formation. Thecultured plant cell gums can also replace prior art gums as a surfacesize which is typically applied after the formation of the sheet atcalender rolls or at the size press. Sandford, P. & Baird, J., supra. Assurface sizes, cultured plant cell gums can impart water resistance, oiland solvent resistance, glue holdout, scuff resistance, physicalstrength, curl control and gloss. The cultured plant cell gums can alsoreplace such prior art polysaccharides as sodium alginate, which is usedas a thickener and dispersant in the pigment coating. The purpose ofsuch an additive is to prevent agglomeration, and to produce adequateflow and leveling of the coating, and to prevent pattern or orange peelin the coating. Sandford, P. & Baird, J., supra.

As exemplified herein, addition of the cultured plant cell gums as abeater aid at the wet end has been observed to result in superiortensile and burst strength, improved resistance to erasure, reduced linton the paper surface and reduced rate of water penetration as comparedpaper manufactured without a beater aid. Without wishing to be bound bytheory, it is believed that at least some of these improvements are dueto a more uniform distribution of pulp fines.

In the adhesives industry, some prior art gums, waxes, tars, and naturalresins have functioned as adhesives when dissolved or dispersed in wateror organic solvents, applied between substrates and thesolution/dispersal allowed to undergo solvent evaporation. Culturedplant cell gums have been found suitable for use in a waterre-moistenable adhesive for paper or aluminum foil sheets. The culturedplant cell gum increases viscosity, thereby moderating the flow duringapplication, and the finished fihn thickness and water retention. Thegum product may also serve as a surface attaching agent. The culturedplant cell gum adhesive, when dried on the surface of paper or aluminumsheets, has good affinity for water and does not cause discoloration ofthe paper or become brittle on aging. The concentration range in theliquid adhesive concentration is between about 1.0 and 3.0% (w/v). Thecultured plant cell gums can be used as an adhesive or cement in otherapplications.

Prior art gums have also been employed in oil and gas field applicationsincluding drilling, well completion (cementing and stimulation) andenhanced oil recovery. As used herein, “oil and gas well fluids” refersto all oil and gas well development or production fluids, includingwithout limitation drilling fluids, cementing fluids, and enhanced oilrecovery injection fluids. Drilling fluids or muds function to transportdrill cuttings to the surface, control formation pressures, maintainbore hole stability, protect productive formations and cool andlubricate the bit and drill string. Prior art gums have been used toimpart greater viscosity to the drilling fluid, to act as suspendingagents for cuttings and weighting materials, and to reduce loss of wateror fluid by preventing penetration into the rock formation. Therheological requirements of the drilling fluid are that it have lowviscosity at high shear rates (i.e., at the drill bit), but highpseudoplasticity to suspend solids in laminar flow. When mud circulationstops, the gel strength is preferably sufficient to suspend solids.Sandford, P. & Baird, J., supra; and Kirk-Othmer Chemical EngineeringEncyclopedia (3rd. ed. 1981) 17:143-166. These rheology requirementshave previously been addressed with combinations of bentonite, celluloseethers, polyacrylamides and xanthan gum. Drilling mud additives forreduction of fluid loss have included carboxymethylcellulose,polyacrylates and xanthan gum. During well cementing, a cement lining isinstalled to isolate the productive zone from the remainder offormations. Fluid loss additives are also used during this stage toprevent cement dehydration and minimize water loss to the formation.Sandford, P. & Baird, J., supra. Following drilling and cementing, acompletion may be used to remove undesirable formation particles anddebris and prevent permeability damage to the producing zone. Completionfluids contain salts for density, and viscosifiers such as xanthan gumto provide suspension for the removal of debris. During wellstimulation, hydraulic fracturing and/or acidizing fluids can be used toenhance hydrocarbon productivity. Hydraulic fracturing fluids requiresuspending agents such as guar or xanthan gums to carry propping solids.Acidizing fluids require a gelling agent effective in high acidconcentrations (e.g., 15% HCl). In enhanced oil recovery, the injectionfluids contain polymers to increase viscosity, resulting in better oildisplacement. Xanthan gum has been a common component in enhanced oilrecovery polymer flooding.

It has now been found that cultured plant cell gums can be employed indrilling fluids to increase viscosity, and as emulsifying, suspending,lubricating agents and fluid loss reduction agents. As an emulsifyingagent in a drilling mud, the cultured plant cell gums can emulsify andstabilize oil-in-water or water-in-oil mixtures. As a suspending agent,the cultured plant cell gums disperse and suspend cuttings and weightingmaterials so as to provide a protective colloid for well equipment. As alubricating agent, cultured plant cell gums can reduce frictionalresistance between the drill string and the formation or casing orduring string raising and lowering. The strong water affinity of the gumproducts can prevent water filtration into surrounding strata duringdrilling or cementing phases. The gum products can also be used asviscosifiers in completion fluids. In hydraulic fracturing fluids, thecultured plant cell gums can be used to impart viscosity, suspendpropping solids and as gelling agents. In enhanced oil recovery,cultured plant cell gums can be used to increase viscosity of theinjection fluid. The concentration of cultured plant cell gum in thedrilling mud, completion, fracturing and enhanced oil recovery injectionfluid is between about 0.1 and 3.0% (w/v). For P. communis gum, a softgel begins to form at about 0.5% (w/v).

For each of the foregoing oil drilling applications, the wholefermentation mixture may be used, i.e., without removal of cells. Thisalternative has the advantage of simplifying the manufacture of oil andgas well fluids. The biodegradability and non-pathogenic nature of thecells makes such alternative possible.

An additional advantage of using cultured plant cell gums in oil and gasfield fluids is that they have much less environmental impact than thoseusing paln oil. This is particularly the case for drilling muds preparedfor off-shore drilling where it is desirable that leakages from the wellbe easily dissipated. Aqueous-based drilling muds dissipate moreeffectively than oil-based muds.

In ink formulations, thickening, suspending and/or emulsifying agentsare used to provide the proper viscosity for application and to increasethe stability of the ink. Lithographic, letterpress and screen printinginks have higher viscosities and frequently contain thickeners.Flexographic (flexo) and rotogravure (gravure) printing inks have lowerviscosities, but use emulsifying or suspending agents for uniformdistribution of the pigment and to prevent the ink from separating.Flexographic inks can be alcohol or water based emulsions. Rotogravureinks also contain an emulsion and have the advantages of excellent,press stability, printing qualities, the absence of fire hazard and theconvenience and economy of water for reduction and cleanup. The inkdistribution systems of flexo and gravure printing presses are simpleand do not provide the means to distribute and level highly viscousinks; therefore, viscosity is typically 5-100 cP. Letterpress and lithoinks can vary in viscosity from under 500 cP for a letterpress-type newsink to over 500 P for special litho ink formulations. In lithography andletter press, uniform and adequate transfer of ink to the printing plateis ensured by a multitude of rollers in the ink distribution unit.Rheology of the litho and letterpress inks is therefore important toroller-to-plate transfer, fidelity in printing, drying speed, holdout,and trapping properties obtained on the substrate. In general, higherpress speeds require lower viscosity inks and slower press speeds employmore viscous inks. Low viscosity ink is used in fine-line flexographyand shallow-cell gravure printing. Printing smooth, dense solids canbest be achieved using higher viscosity ink. Rheology is also importantas a color strength determinant. Over-pigmentation leads to a morethixotropic ink, thereby creating a balancing relationship between colorintensity and rheology. Kirk-Othmer Chemical Engineering Encyclopedia,supra, Vol. 13, pp.374-376. Lasday, S. (ed.) Handbook for GraphicCommunications: (1972) Ink, Paper, Binding, Vol 6., pp. 6-13.

It has been found that cultured plant cell gums can be used asemulsifying, suspending and/or thickening agents in a variety ofprinting inks, including litho, letterpress, screen printing,flexographic and gravure inks. The gum concentration in flexo inks canbe between about 0.5 and 4.0% (w/v).

Additionally, in offset lithography, prior art gums have been used asemulsifying agents and viscosifiers in lithography solutions. Offsetlithography is a planographic process where the image and non-image arein the same plane. The image area is oil receptive and the non-imagearea is water receptive so that following wetting of the plate with thefountain solution, the ink, when rolled across the plate will only beattracted to the oil receptive areas. As used herein, “lithographysolution” refers to any non-ink solution used in lithography, includingfountain solutions, sensitizing solutions and protecting solutions. Thefountain solution is a desensitizing solution which prevents ink fromadhering to the plate. Fountain solutions have contained gum arabic,typically at 0.2% (w/w). Lasday, S., sgpra, Vol. 6, pp. 93-95. Thedesensitizing use of gum arabic has taken advantage of the goodwettability imparted to the fountain solution and also of the viscositycontrol that allows the wash solution to cling to the plate withoutrunning off or forming isolated droplets or pools on the plate. On metalplates, the desensitizing effect might be caused by the formation of aninsoluble film of EG Aluminum or Zinc Arabate. A more plausibleexplanation is that the film of gum is absorbed by the plate. Studieshave shown that such films occur on plates of zinc, aluminum, copper,silver, iron tin, lead, glass and fused silica. These films are notmono-molecular but are composed of many molecular layers. LSC PrintingInks, Reinhold Publishing Corporation, New York (1940) pp. 230, 334,346, 398-9 and 417. Measurement of the wettability of the desensitizingsolutions can be evaluated by measurement and study of the contactangles. In this process a section of the plate is partially immersed inwater or in a solution of the gum to be tested. The plate is then turnedat an angle to the surface of the liquid until the meniscus appears tobe eliminated. The resulting angle of the plate to the surface of theliquid is known as the contact angle and is the measure of thewettability of that particular plate with the solution being tested.Read REF Modem Lithography (1951)47:62.

Cultured plant cell gums can be used as emulsifying agents insensitizing or fountain a solutions for the plates during operation andin protecting solutions during storage. The concentration range of thegum in fountain solutions is between about 0.01 and 2.0% (w/w).

In the textile industry, gums have been used as sizing and thickeningagents. Sizing agents act during textile manufacture by binding theloose fibers of the warp, thereby imparting strength, flexibility andsmoothness to the warp, allowing weaving to proceed efficiently.Thickeners control the viscosity of various formulations used in thetextile industry including dyes, printing inks, coating and flockingsolutions. Prior art gums, including guar, algin and xanthan gums havebeen used in printing and dyeing solutions. Sandford, P. & Baird, J.,supra. Cultured plant cell gums can be useful as sizing or thickeningagents in the textile industry. As exemplified herein, the gum productcan function as a thickening agent for dyestuff used in wool and cottonfabric printing. The concentration range of the gum product in thedyestuff is between about 0.1 and 5.0% (w/v). Modified approaches can beused in the reactive dyestuff process and direct vat dyestuffs for silkand hydrophobic man-made fibers (nylon, acrylics, polyester andacetates).

In the paint industry, viscosifiers, thickeners, emulsifying agents,suspending agents, and dispersants are used to improve flow propertiesof the paint so that a smooth coat of desired thickness can be appliedto a vertical surface without sagging, and to stabilize the paint bypreventing coagulation and pigment settling. Thixotropic character ofthe paint is important in providing good levelling, prevention ofrunning, and avoidance of segregation or stratification of the paintduring storage. Sandford, P. & Baird, J., supra; and Gamble, D. & Grady,D., U.S. Pat. No. 2,135,936 (1938). As exemplified herein, culturedplant cell gums can be used as emulsifying agents in an acrylic resinpaint or an oil emulsion paint. The concentration range of the gumproduct in acrylic or oil based paint is between about 0.2 and 0.3%(w/v).

In ceramics manufacturing, a glaze or a colored, opaque or transparentcoating is applied to the ceramics before firing. The glaze forms ahard, nonporous surface. Glazes are usually made from powdered glasscombined with colored oxides of such elements as cobalt, chrome,manganese or nickel. The mixture of powders is suspended in water andapplied to the ceramic surface by spraying brushing or dipping. Theglaze is then dried and fixed onto the ceramic surface by firing.Emulsifying agents, suspending agents or dispersants can be used touniformly distribute the pigments in the glaze. The glaze causes thepigment to adhere to the surface during firing. As exemplified herein,cultured plant cell gums can be used as an emulsifying and suspendingagent to produce a glaze of superior consistency, clarity and stability.Further, it has been found that if BLM (Brewers Liquid Maltose) is usedas a carbon source during culturing of N. plumbaginifolia, the recoveredgum product imparts excellent film-forming properties to the glaze. Thegum product concentration range in the liquid glaze is between about0.05 to 3.0% (w/v).

Cultured plant cell gums can also be useful in ceramics forming byplastic extrusion. Completely nonplastic materials can be extruded withthe addition of suitable plasticizers such as gums, starches,polyvinylalcohol, waxes and wax emulsions. Grayson, M. (ed.) Kirk-OthmerConcise Encyclopedia of Chemical Technology (1985) p. 237. Culturedplant cell gums can replace prior art gums in such processes. Inceramics forming by slip casting, cultured plant cell gums can be usedin the suspension of raw materials to ensure uniform dispersion of theclay and other solid particles in the water.

In cleaning detersive systems, absorption of bath components to thesubstrate surface may be the most important and fundamental detergencyeffect. Adsorption is the mechanism whereby the interfacial free energyvalues between the bath and the solid components (substrate and soilthereon) of the system are lowered, thereby increasing the tendency ofthe bath to separate the solid components from one another. Surfactantadsorption reduces soil-substrate interactions and facilitates soilremoval. Kirk-Othmer Chemical Engineering Encyclopedia, supra, Vol. 22,p. 408.

In cleaning detergent manufacturing, the addition of materials toincrease viscosity and film-forming properties can enhance surfactantand substrate surface interactions, particularly for vertical surfaces.As exemplified herein, cultured plant cell gums have been found to beuseful in improving the viscosity and film-forming properties ofdetergents. In particular, it has been found that use of BLM as a carbonsource in the culturing of N. plumbaginifolia produces a gum productthat can impart improved film-forming properties to the cleaningdetergent. This is particularly useful for cleaning detergents used toclean vertical surfaces. Detergents can also contain soilantiredeposition or suspending agents, such as carboxymethylcellulose,polyvinylalcohol and polyvinylpyrollidone. These antiredeposition agentsare believed to function by absorbing onto either the substrate or thesoil particle, and imparting electrical charges that reduce the affinitybetween the soil and substrate. Sandford, P. & Baird, J., supra. It isbelieved that cultured plant cell gums can also function as anantiredeposition agent by coating the substrate and/or soil particles.The gum product concentration range in cleaning detergents is betweenabout 1 and 10% (w/v).

Plant cell gums can function as encapsulating agents. As such, the plantcell gum can be used for encapsulation of lemon oil (or other suitableoils) in detergent powders and other dry or powdered cleaning agents.

Plant cell gums have been found useful in the manufacture of chemicalcompositions with decreased tendency to form fine mists or satellitedroplets. Of particular interest are applications in agriculture toprevent unwanted dispersion of potentially harmful agriculturalchemicals (insecticides, fungicides, etc.) on spray application.

Cosmetic lotions and creams are water-in-oil or oil-in-water emulsionsemploying emulsifying and stabilizing agents. Emulsifiers, being surfaceactive agents, lower surface and interfacial tensions and increase thetendency of the lotion or cream to spread. A purified acidicheteropolysaccharides obtained from cultured Polianthus has been used incosmetic creams, lotions, shampoos and cleansing foams. Otsuji, K. etal. EP 0 285 829, published Oct. 12, 1988. As exemplified herein,cultured plant cell gums can be used without prior purification of gumfractions in cosmetic lotions and creams. The gum product concentrationrange in the cosmetic lotions and creams is between about 0.5 and 4.0%(w/w). Plant cell gums have been found useful for the formation of softgels which spread well on the skin and feel smooth and supple to thetouch. Humectant and perfume can optionally be added to these soft gelsto provide moisturizing cosmetic gels. Plant cell gums have also befound useful in the preparation of deodorants, hair styling gels, andshampoos and conditioners.

Other applications for cultured plant cell gums include thickeners,emulsifiers or suspending agents for photographic preparations;thickeners for explosives; thickeners and suspending agents for foundrywash coats; thickeners, foam stabilizers and film formers forfire-fighting fluids; emulsifiers and suspending agents for flowablepesticides, suspension fertilizers and animal liquid feed supplements.

The advantages of the cultured plant cell gums over prior art gumsinclude lower production costs, improved purity and improved productionreliability. Because the production of cultured plant cell gums does notrely on labor-intensive harvesting of gum exudate from trees (e.g., asis required for gum arabic) or harvesting of seeds or plants forextraction (e.g., guar gum, agar algin, or carrageenan), and can insteadbe produced under automated conditions, labor costs associated with theproduction of cultured plant cell gums can be lower. As discussedhereinabove, in agar production, the harvesting of red seaweed is laborintensive in that it is carried out by hand; in some areas of the world,divers in full pressure suits collect individual plants in deep water;in other places, the seaweed can be collected at low tide without theuse of diving equipment. Carrageenan or Irish Moss is produced fromanother red seaweed harvested by raking and hand gathering. Algin isproduced from brown algae which can be harvested manually or with smallmechanical harvesters. Sandford, P. & Baird, J. (1983) in ThePolysaccharides, Academic Press, Inc. Vol. 2, pp. 411-491. Additionally,since production of cultured plant cell gums is carried out infermentation facilities, production does not rely on weather and istherefore more reliable than prior art gum production. See, for example,Meer et al. (1975) Food Technology 29:22-30. Further, because culturedplant cell gums are produced in fermentation facilities, they can bepurer than prior art gums. As discussed hereinabove, because gum arabicis hand collected, it is seldom pure; samples are classified accordingto grade which depends on color, contamination with foreign bodies suchas wood or bark (VanNostrand's Scientific Encyclopedia, supra at p.1389).

An advantage of cultured plant cell gums over xanthan gum produced bycultured Xanthamonas campestris is that the cultured plant cells do notpose the same cell disposal problem presented by X. campestris, a plantpathogen (Scaad, N. W. (1982) Plant Disease 66(10):882-890). Further,cultured plant cell gums are less expensive than xanthan gum for avariety of applications, including drilling fluids (e.g., Kirk-OthmerChemical Engineering Encyclopedia (3rd. ed. 1981) 17:153).

A further advantage of the subject gum product is that it can often beused in smaller quantities than prior art gums to achieve comparableeffectiveness as an emulsifying, is stabilization, suspending,thickening, or gelling agent, as a film forming or coating agent, or asa protective colloid.

All references cited are incorporated herein by reference in theirentirety.

The following examples are provided for illustrative purposes only andare not intended to limit the scope of the invention.

EXAMPLES Example 1 Establishing Suspension Cultures

1.A.—Phleum pratense

Seeds var. Kahu from Hodder & Tolley, seed merchants, 17 Binney Rd.Marayong, Australia, were sterilized by rinsing in ethanol and thensoaking 5 minutes in hypochlorite (“chlorize” 1:4). The seeds were thenrinsed three times with water and transferred to either liquid or solidmedium of Hale et al., supra containing 2 mg/l 2,4-D.

Suspension cultures were initiated from seeds germinating on eitherliquid culture or callus culture. In the liquid culture, most seedsgerminated after five days. The seed and liquid were chopped in a smallsterile blender and then returned to an Erlenmeyer flask and shaken fora further two weeks. The resulting culture was propagated by regularsubculturing every 2-3 weeks into suspension culture.

The seeds germinating on agar medium began to form callus immediately.The small calli were dissected off and transferred to fresh agar medium.The calli were subcultured every 3-4 weeks. Initially, the calli aremucoid, but after a number of subcultures, they lose their mucoidappearance. Suspension cultures initiated from mucoid calli produced 2-5g/l of polysaccharide. Suspension cultures initiated from calli thatlost their mucoid appearance and no longer produced polysaccharide.

The suspension medium and procedure were those employed in Hale, A. etal., supra.

Within three days of initiation into the suspension medium, culturefiltrates are extremely viscous (i.e., filtrate runs from a 5 mL bulbpipette in about 70-80 seconds, as compared to 15 seconds for water and16-20 seconds for Pyrus cell culture filtrate). Also, there is verylittle growth of cells, so the filtrate volume on harvesting isvirtually the same as the culture volume (i.e., the packed cell volumeis negligible). While polysaccharide production is lost from callus andsuspension cultures on repeated subculture, this does not create aproblem as it is easy to initiate a new cell line.

1.B.—N. plumbaginifolia

Callus was initiated from seeds cultured on 20-30 mL CSV (Gibson et al.(1976) Planta 128:233-239; and Schenk, R. & Hildebrandt, A. (1972) Can.J. Bot. 50:199-204)) medium (below) solidified with 0.5% (w/w) agar. Thecallus was maintained on the same solid medium, in the dark at 27° C.Maintenance subculturing occurred approximately every 3 weeks. If dryingor discoloration of the culture was observed, it was immediatelysubcultured.

All stock solutions were made up with Milli-Q™ water in glass bottles.

CS Macro salts NH₄NO₃ 24.8 g  KNO₃ 50.1 g  (NH₄)H₂PO₄ 9.2 g CaCl₂ · 2H₂O4.0 g MgSO₄ · 7H₂O 8.0 g

The solution was made up to 1 liter with Mili-Q™ water and stored at 1°C. in glass

CS organics Thiamine-HCl 100 mg Nicotinic acid 1000 mg  Pyridoxine-HCl100 mg

The solution was made up to 200 mL with Milli-Q™ water and stored at−20° C. in glass bottles.

CS micro salts MnSO₄ · 4H₂O 6.5 g   H₂BO₃ 2.5 g   ZnSO₄ · 7H₂O 0.5 g  KI 0.5 g   CuSO₄ · 5H₂O 100 mg  NaMoO₄ · 2H₂O 50 mg CoCl₂ · 6H₂O 50 mg

The solution was made up to 500 mL with Milli-Q™ water and stored at−20° C. in glass bottles.

CS Iron solution Na₂EDTA · 2H₂O 2.0 g FeSo₄ · 7H₂O 1.5 g

The EDTA was dissolved in 60 mL Milli-Q water, while stirring andheating. It was then cooled to room temperature and the FeSO₄.7H₂O wasslowly added while also adding NaOH (10 M=400 g/liter) to keep pH at5.9. The solution was made up to 100 mL with water and stored at −20° C.in glass bottles.

To prepare one liter of CSV medium, the stock solutions and solids weremixed in the following quantities in approximately 800 mL of Milli-Q™water:

CS Macro 50 mL CS Micro 1 mL CS Iron 1 mL CS Organics 1 mL Sucrose 30 gmyo Inositol 1 g

The pH was adjusted to 5.8 (20-30 drops of 1M KOH). This medium can bemodified in various ways without adverse effect, e.g., inositol can bereduced or deleted. The hormone stocks were added in the followingquantities:

2.0 mL 2,4-D (stock 1.0 mg/mL)

0.5 mL of kinetin (stock 0.1 mg/mL).

The solution was then made up to 1 liter with Milli-Q™ water andsterilized for 20 minutes at 10 psi (116° C.).

Suspension cultures were passaged into fresh CSV medium at 7-dayintervals using a 10% inoculum (i.e., 2 mL into 20 mL, 20 mL into 200mL). Suspension cultures were maintained at a 27° C. at a shaker speedof 100 rpm. The cultures were monitored visually for departures fromnormal color and cell growth patterns. Cultures were also monitored forsterility (i.e., contaminating organisms) and healthy cell morphology(e.g., cell stress).

1.C.—Pyrus communis(Green Pear)

Callus was initiated from fruit cultured on 20-30 mL pear BAL (balanced)medium (below) solidified with 0.5% (w/w) agar. The callus wasmaintained on the same solid medium, in the dark at 27° C. Maintenancesubculturing occurred approximately every 4 weeks. If drying ordiscoloration of the culture was observed, it was immediatelysubcultured.

All stock solutions for the pear BAL media were made up using Milli-Q™water in glass bottles. Vitamins and hormone solutions were stored at−20° C.; all other solutions were stored at 1° C.

Macro elements NH₄NO₃ 165 g KNO₃ 190 g MgSO₄ · 7H₂O  37 g

The Macro solution was up to 1 liter with water.

Micro elements H₃BO₃ 1 g ZnSO4 · 7H₂O 1 g MnSO₄ · H₂O 1.44 g NaMoO₄ ·2H₂O 0.029 g CuSO₄ · 5H₂O 0.0025 g (*) CoCl₂ · 6H₂O 0.0025 g (*)

The Micro solution was made up to 100 mL with water. (*) To obtain 2.5mg of these salts, 25 mg of each was weighed out in separate containers,and dissolved in 10 mL Milli-Q™; 1 mL of each solution was then used.

Vitamins Ca pantothenate 0.1 g myo-Inositol 10.0 g Biotin 0.001 g (*)Nicotinic acid 0.001 g (*) Thiamine-HCl 0.1 g Pyridoxine-HCl 0.05 g

The vitamin solution was made up to 100 mL with water. (*) A stocksolution containing 1 mg of Biotin+1 mg of Nicotinic acid per 10 mL wasprepared as follows: 10 mg of both vitamins was dissolved in 100 mL ofMilli-Q; 10 mL of this solution was used to make up 100 mL of StockVitamins.

KH₂PO₄ (Potassium Dihydrogen Orthophosphate)

KH₂PO₄ 17 g

The solution was made up to 1 liter with water.

CaCl₂.2H₂O (Calcium Chloride Dihydrate)

CaCl₂ · 2H_(2O) 6 g

The solution was made up to 100 mL with water.

Fe · EDTA FeSO₄ · 7H₂O 6.86 g Na₂ EDTA · 2H₂O 9.17 g

The EDTA was dissolved in 1 liter of Milli-Q™ (magnetic stirrer, roomtemperature). The ferrous sulphate was dissolved in the EDTA solution.The resulting solution was brought to a boil, cooled and stored in screwcapped glass bottle at 1° C.

KI 0.03 g

The KI was dissolved in 20 mL Milli-Q.

2.4-D (2,4-dichlorophenoxyacetic acid) 0.1 mg/mL

2,4-D 50 mg

The 2,4-D was dissolved in 5 mL of commercial grade ethyl alcohol (95%).The 2,4-D was injected slowly under the surface of 495 mL of Milli-Q™water, using a Pasteur pipette and a magnetic stirrer.

To make up the pear BAL medium, the concentrated stock solutions andsolids were mixed (magnetic stirrer) in the quantities indicated belowand water added to approximate 900 mL.

Macro elements 10 mL Micro elements 1 mL Vitamins 1 mL KH₂PO₄ 10 mLCaCl₂ 2.5 mL Fe.EDTA 2.5 mL KI 0.5 mL 2,4-D 10 mL L-Asparagine 180 mgL-Ascorbic acid 50 mg Thiourea 25 mg Sucrose 40 grams

The pH was adjusted to 5.8-6.0 with KOH (0.1 or 1M). The final volumewas adjusted to 1 liter with water. For solid medium, 0.5% (5 g/liter)agar was added after adjusting pH and volume. The final medium wassterilized for 20 minutes at 10 psi (116° C.).

Suspension cultures were passaged into fresh BAL medium at 14 dayintervals using a 20% inoculum. The cultured were maintained at 27° C.at a shaker speed of 100 rpm. Cultures were monitored for sterility,cell morphology, and departures from normal culture color and cellgrowth. After subculturing into fresh BAL medium, the packed cell volume(PCV) of the old culture is measured to assess whether the cultureconditions are successfully maintaining the cell line in a stable growthpattern. If the PCV declined progressively over several subcultures, thecell line was revived with a single passage on double phosphate pear BALmedium.

1.D—Enhanced Polysaccharide Production Using BLM

When BLM was used as a carbon source to enhance polysaccharideproduction by Nicotiana or Pyrus, it was typically used at a culturemedium concentration of between about 80 to 200 g/liter of medium, orpreferably at about 162 g (wet weight) per liter of medium.

Example 2 Recovery of Gum Product from Cultured N. plumbapinifolia

N. plumbaginifolia whole broth was harvested from a fermentor. The wholebroth was filtered using a filter having a pore size of about 100 μm.The filtrate was then heated to 80° C. for 30-60 minutes to denatureenzymes in the filtrate. The filtrate was then cooled. Complexant (e.g.,Na₂EDTA.2H₂O; 1 g/l) was added either prior to filtration, afterfiltration and prior to heating, or after cooling.

In some cases, the filtrate was stored prior to further processing. Whenstorage time was longer than 18 hours, preservatives, e.g., 1.0 g/lpotassium sorbate and 0.34 g/l sodium metabisulfate, were added. Thesepreservatives allowed storage at ambient temperatures (15-25° C.) insealed containers for prolonged periods.

The filtrate, warmed to 30-80° C. to reduce viscosity, was nextconcentrated by ultrafiltration (10,000 MW membrane, Amicon ModelDC10LA) to about 20-25% of its original volume or until viscosity madefurther significant concentration difficult. The concentrate was thendiafiltered using the same membrane with five equal volumes of distilledH₂O, and concentrated again by ultrafiltration to the point at whichviscosity or gelling inhibited further progress.

Where the gum product was intended to be used in industrial compositionssuch as in drilling mud, adhesives, cleaning detergents, dyestuffs,paper, acrylic resin and oil emulsion paints, or printing ink, theconcentrate was directly spray dried (Niro Production Minor, NiroAtomizers, Denmark) using a 200° C. inlet temperature and a 100° C.outlet temperature.

Where the gum product was intended to be used in foods, pharmaceuticals,or cosmetics, the concentrate was further purified by an alcoholprecipitation method comprising a precipitation and washing step. Theconcentrate was chilled to 1-4° C., and NaCl or KCl was added as aconcentrated solution, followed by slow addition with stirring of 2-4volumes cold (1-4° C.) ethyl or isopropyl alcohol. The NaCl or KCl wasadded in an amount to give a concentration of 0.03-0.1% w/v in thealcohol-containing mixture. The mixture was allowed to stand at 1-4° C.for 1-18 hours and then filtered using 2-4 layers of surgical gauze. Thefiltrate was washed in 67-80% alcohol at 1-4° C. and the wash wasremoved by filtration using 2-4 layers of surgical gauze. The alcoholcan be recovered and recycled by distillation.

Where further purification was desired, the alcohol purificationprocedure was repeated one or more times. A variation of thepurification procedure comprises repeated precipitation and filtrationsteps without intervening washing steps.

The purified material was then directly drum dried (Blaw-Knox Co.Buffalo, N.Y.). Alternative drying methods are fluidized bed, vacuumtumble drying and “flash-spin” drying. The purified material can also bespray-dried or freeze-dried if first rehydrated with 1-2 volumesdistilled H₂O.

Example 3 Functional Assessment of Recovered Gum Products

3.A.—Emulsion Testing: Measurement of Droplet Size, Turbidity andStability

A comparison of the emulsifying properties of Pyrus gum and a prior artgum, gum arabic, was conducted to determine whether the claimed gum hasemulsifying qualities comparable or improved relative to the prior artgum. Aqueous solutions of Pyrus gum and gum arabic were mixed withD-limonene oil to produce emulsions, which were then tested for dropletsize, turbidity and shelf life stability.

In order to clarify or reduce complexing of the pectic fraction of thePyrus gum prior to use, 5 grams of Pyrus gum were dissolved in 500 mL ofdistilled water and boiled for 5 minutes. Concentrated EDTA solution wasadded until the insoluble pectic material was dissolved. The solutionwas filtered through two layers of Whatman glass fiber filter paper GF/Funder vacuum and dialyzed (MW cutoff 14,000-16,000) against distilledwater at 4° C. for 24 hours. The volume of the solution was then reducedunder vacuum by rotary evaporation and freeze dried. Gum arabic wasobtained from Sigma, No. G-9752. D-Limonene (p-mentha-1,8-diene) wasobtained from Bush Boake and Allen.

Stock solutions of gum arabic (250 mg/mL) and Pyrus gum (62.5 mg/mL or12.5 mg/mL) were pipetted in duplicate to give final concentrations of0, 0.2, 0.5, 1, 5, 10 and 20% (w/v). The Pyrus solution could not beprepared at concentrations greater than 5% (w/v) due to its viscosityand gelling properties. Twenty percent D-limonene oil in water emulsionswere prepared by injecting the oil into the aqueous solutions under thesurface of the solutions while being mixed in an Ultraturrax (YstalT1500, 25-240V, West Germany) at setting 4 for 15 seconds. The speed ofthe Ultraturrax was increased to setting 7 for 45 seconds to produce thecloud emulsion. The emulsions were allowed to stand for 0.5 hours toallow bubble dispersal.

To determine emulsion capacity, droplet size, turbidity and shelf-lifewere measured for each emulsion. Emulsion capacity increases withdecreased droplet size, increased turbidity and increased shelf-lifestability. The droplet size of the cloud emulsion was examinedmicroscopically by placing 2 drops of the emulsion on a slide anddiluting with 2 drops of water and estimating droplet size using acalibrated eye piece graticule. Cloud turbidity was measured by dilutingduplicate 5 μl aliquots of cloud emulsion into 5 mnL of 0.1% (w/v)sodium dodecylsulphate and measuring absorbance at 500 nm. Cloudemulsions were tested for shelf-life stability by centrifuging at 2,500rpm for 10 minutes and observation of the resulting separation of oiland water phases. The results are set forth in Table 2.

From these results, it is seen that when emulsifying 26% D-limonene inwater, Pyrus gum on a weight for weight basis produces smaller dropletsat a lower concentration than gum arabic. For example, at 0.2% (w/v) ofPyrus gum, the emulsion mixture has a film of free oil, a cream layerstable to centrifugation, oil droplets of 1-20 gm and a cloud turbidityat 500 nm of 0.127. In contrast, 0.2% (w/v) gum arabic in an emulsionmixture has an unstable cream which separates completely to oil oncentrifugation, has a larger droplet size (10-20 μm) and an averagecloud turbidity reading of 0.023 at 500 nm. These results indicate thatthe Pyrus gum has improved emulsifying qualities relative to those ofgum arabic at the same concentration.

Emulsion stability can also be assessed by the following method. Anoil-in-water emulsion was produced with a range of gum productconcentrations (e.g., 0.2, 0.5 and 0.7% (w/v)):

gum (g) 0.1 0.25 0.35 oil (mL) 10.00 10.00 10.00 water (mL) 40.00 40.0040.00 Total 50.00 50.00 40.00

The gum product was dissolved in water using the ultraturrax (JohnMorris Scientific Equipment) at a setting of 4. Oil (Crisco,polyunsaturated blend) was then added while mixing and held at setting 4for 45 seconds. The solution was further mixed at setting 8 for 45seconds. The emulsion obtained was poured into 50 mL measuring cylinders(21 mm internal diameter), sealed with aluminum foil and stored at 27°C. It was then observed for up to a week. Creaming or separation wasexpressed in percentage volume.

The volume of oil can be varied to provide an HLB in the emulsion thatis typical for the intended application.

For measuring the stability of a water in oil emulsion comprising acultured plant cell gum, ASTM method 3707 or an adaptation thereof canbe used.

3.B.—Viscosity Testing

The flow behavior of the gum product in aqueous solutions or mixtureswas assessed over a range of gum concentrations, temperatures and shearrates. The gum product was dissolved in water using the ultraturrax atsetting number 4. The solution was then stirred and heated to 60° C.Viscosity was measured at decreasing temperatures from 60° to 10° C.using an Epprecht Rheometer (Contraves AG Zurich) at various shearrates. Results were plotted as viscosity versus temperature fordifferent shear rates.

Viscosity plotted as a function of shear rate indicates the thixotropicnature of the gum. Thixotropic profiles indicate whether a gum is suitedfor particular applications where shear thinning is required (e.g., indrilling muds) at the drill bit.

Viscosity plotted as a function of temperature indicates the suitabilityof the gum as a viscosifier or thickener over the operating temperaturerange of the intended application.

3.C.—Gel Strength Testing

Gel strength is assessed by measuring rupture strength, shear modulusand back extrusion. Back extrusion is of particular interest because itcan distinguish between and characterize soft gels and viscous fluids.

3.C.1—Rupture Strength

Rupture strength is the force required to compress and rupture a gelsample. For rupture strength, the force is proportional to the sampleweight.

The gel samples were prepared by mixing P. communis gum (0.2-0.5% (w/v))or N. plumbaginofolia (0.5-1.0% (w/v)) in water in 50 mm plastic petridishes and storing them at 15° C. overnight. Rupture strength wasmeasured by compression on the Instron 1122, using a probe of 150 mm indiameter at a cross-head speed of 50 mm/min.

3.C.2—Shear Modulus

Shear modulus is a measure of the force required to shear/cut the gel.Shear modulus is expressed as stress divided by strain. For shearmodulus, the force is proportional to the sample weight.

The gel samples were prepared as in C.1 in a 24 mm diameter glass vialand stored at 15° C. overnight. Shear force was measured on a modifiedpuncture strength meter (Oakenfiill, D. G. et al. (1987) “A method fordetermining the absolute shear modulus of a gel from a compression test”in Gums and Stabilizers for the Food Industry, Vol 4, Phillips, G.O. etal. (eds.) IRL Press, Oxford) with a probe of 3 mm in diameter at thecross head speed of 5 mm/min for 20 seconds. Shear modulus was thencalculated using a mathematical model set forth in Oakenfull, D. G. etal. (1987).

3.C.3—Back-extrusion

Back-extrusion force is the force required to compress and shear a gelsample. In back-extrusion, force is independent of sample weight.

The gel samples were prepared as in C.1 in 200 mL beakers of 64 mm andstored at 15° C. overnight. Back-extrusion was performed on the Instron1122 by plunging a probe of 60 mm in diameter at a speed of 100 mm/minto a depth of 50% into the gel.

3.C.4.—Effective Temperature Range of Gel: Determination of Melting andSetting Points

The melting point is determined by observing the temperature at which a10 mL gel begins to melt in a 11 mm diameter spectrophotometric tube.The determination was aided by observing small glass beads (0.08 g)sinking into the melting gel. As there can be temperature gradientswithin the gel, a melting range can be observed. The experiment wascarried out in a Thermoline waterbath in 5° C. steps.

The setting point was determined by observation of gelling inspectrophotometer tubes. The gel samples in the tubes were storedovernight (18 hours) at a range of temperatures, and the tubes were theninverted to observe if setting had occurred. The temperatures testedwere 6°, 10°, 15°, 20°, 25°, 27°, 30°, 37.2° and 45° C.

3.D.—Encapsulating Capacity

Encapsulating capacity can be assessed by evaluating a gum-containingspray dried emulsion in terms of flow characteristics and stability, asdescribed in Example 3 of a U.S. Pat. Nos. 5,296,245 or 5,133,979.

3.E.—Adhesive Capacity

Adhesive capacity can be measured by using standard methods such as ASTM(American Society for Testing Materials) method D1713 (“BondingPermanancy of Water- or Solvent-Soluble Liquid Adhesives for AutomaticMachine Sealing Top Flaps of Fiberboard Specimens”) and D1581 (“BondingPermanancy of Water- or Solvent-Soluble Liquid Adhesives for LabellingGlass Bottles”), or adaptations thereof.

Example 4 Papermaking—Preparation of Paper Hand Sheets

A superior strength paper can be produced using the procedure describedin Australian Standard 1301 APPITA P203s/80 by adding N. plumbaginifoliagum product at the wet end to improve the physical properties of the drysheet. The observed improvements include increased paper strength (bothburst and tensile), greater resistance to erasure, reduced “fuzz” orlint on the paper surface and reduced rate of water penetration ascompared to hand sheets prepared without a gum beater aid. The gumproduct allows for a retention of wet strength and improved yield byproviding a more uniform distribution of fines.

The following method (Australian Standard 1301 APPITA P203s/80) was usedfor the preparation of hand sheets:

Commencing with wood fibre pulp (sourced as chemically treated pulp,semi-chemical pulp, or mechanical pulp or recycled pulp), the gumproduct was dissolved in a quantity of water sufficient to produce a 2%solids solution. One liter of the dissolved solution was added to 4liters of pulp placed in a container. Adequate mixing was ensured bysparging for at least 15 minutes. A sample of 500 mL was then place intoa larger tapering 15 liter vessel with a 60 mesh screen at the base 100mm in diameter. A further 10 liters of processed water was added and themixture was sparged from the base of the vessel for at least 15 secondsto ensure thorough mixing. The base valve was then opened, allowingprocessed water to drain away, retaining all of the fibers on the wiremesh screen. The base screen was removed from the unit base and coveredwith a blotter, allowing the wet fibrous mat to be retained by theblotter. Successive cycles produce a number of samples which are thenstacked and pressed in a stack to remove excess water. They were thenplaced in a drying cabinet and maintained at a standard 23° C., 50%relative humidity until testing.

Testing revealed that the subject gum-containing paper has superiortensile strength, stretch, work to rupture and extensional stiffness onan Alweitron Universal Testing machine. Methods for testing paper areknown in the art and include, e.g., Australian Standard 1301.403s-89 for“Bursting Strength of Paper;” Australian Standard Appita P404s-81 for“Tensile Strength of Paper and Paperboard;” Australian Standard1301.419s-89 for “Water Vapour Transmission Rate of Paper;” AustralianStandard 1301.411s-89 for “Water Absorptiveness of Paper and Paperboard(Cobb Test);” and Australian Standard Appita P406m-86 for “BendingQuality of Paperboard.”

Example 5 Adhesives—Preparation of Re-moistenable Adhesive

A satisfactory adhesive for envelopes, labels, stamps and aluminum foilsheets, which is of the water re-moistenable type, was prepared asfollows:

1. N. plumbaginifolia (BLM carbon source) 1000 g 2. Sodium Chloride 20.5g 3. Glycerol 20.5 g 4. Potato starch 20 g 5. Water 1300 mL 6.Preservative 1 g

The water was placed in a high speed mixer and mixing was begun at aslow speed. The gum product was slowly added, allowing it to fullydissolve in the mixing process. After 4 minutes, the sodium chloride,glycerol, starch and preservative were added. After thorough mixing, themixture was left to stand for 1½ hours.

This produced an adhesive which was applied to the surface of paper anddried. It remained inactive until moisture was reapplied. It was foundto be a superior gum for use in these applications as it has goodaffinity for water and does not cause discoloration of the paper orbecome brittle on aging. It was found that the adhesive glued pieces ofaluminum foil to paper very firmly and also glued pieces of papertogether in a manner similar to commercial adhesive pastes.

Example 6 Oil and Gas Well Applications—Preparation of Drilling Mud

A satisfactory drilling mud or fluid can be prepared in stirred tanks asfollows:

A large 1,000 liter tank was filled with water. About 6% by weightbentonite (montmorillonite) was added while stirring slowly andcontinuously until dissolved. In a second 1,000 liter tank filled withwater, about 3% by weight N. plumbaginifolia gum product was added whilestirring slowly until dissolved. In a third holding tank, equalquantities of gum product mixture and bentonite mixture were mixed. Thisproduced a basic drilling fluid to which was added up to 30% solids ofbarium sulphate or 30% chalk as weighting agents depending upon thenature of the surrounding rock structure. If desired, a biocide can beadded to prevent fermentation during storage or down-hole.

The resulting drilling fluid has increased viscosity, and can provide animproved flow of material from the bit to the surface and a uniformdispersion of the solids, thereby acting as a protective colloid. It canalso lubricate and reduce fluid loss into porous rock. The resultingdrilling fluid is particularly efficacious in providing a uniformsuspension and maintaining a consistent fluid in drilling through shalelayers, broken rock that has been stabilized, or magnesium or calciumcontaining rock. During cementing, a stabilizing fluid containing thesubject gum product will also have reduced fluid loss.

The N. plumbaginifolia gum product, when in an aqueous dispersion withcalcium, possesses gelling properties. Such gelling properties canenhance suspension of solids in a drilling mud even when flow hasstopped.

Example 7 Printing Applications

7.A. Preparation of Printing Ink

A satisfactory emulsion or suspension water-based flexo ink for printingwas prepared using the N. plumbaginifolia gum product as a suspensionagent to provide uniform dispersion of the pigment elements and preventthe ink from separating. To a typical ink formulation of:

1. Carbon Black

2. Mineral Oil

3. Sodium silicate

4. Sodium carbonate

5. Water

was added about 2% by weight of gum product to produce a fine uniformstable suspension of the solid ingredients. Using a high speed mixerrunning at low speeds the gum product was added to the mixture untilthoroughly dispersed. The emulsion mixture was left to stand for 1½hours prior to use.

7.B. Preparation of a Lithography Fountain Solution

The N. plumbaginifolia gum product provides a satisfactory substitutefor gum arabic in several lithography solutions or mixtures includingthe plate sensitizer solution, the fountain solution and the protectingsolution (used during plate storage). The gum product imparts goodwettability particularly to the fountain solution. It also supplies theviscosity required to allow the fountain solution to cling to the platewithout running off or forming isolated droplets or pools on the plate.

A fountain solution was prepared as follows:

1. Water 700 mL 2. Propylene Glycol 50 mL 3. Biocide Parabens(methyl/ethyl- 1 mL hydroxy parabenzoic acid at 0.5-2.0% (w/v) in wateradjusted to pH 7.0 with phosphate buffer 4. gum product solution 3%(w/w) 200 mL 5. pH buffer 40 mL

All ingredients other than the gum product solution were added to amixture, and stirred until dispersed (10 mins). The gum product solutionwas then added, and stirring was continued. The mixture was then allowedto stand for 30 minutes before use.

7.C. Comparison of N. plumbaginifolia Gum to Gum Aabic in FountainSolutions

The following formulae were made up by Cetec Pty. Ltd., a consultant.All values (except pH) are w/w percent.

F1 F2 F3 F4 Water 70 90 70 90 Propylene Glycol 5 — 5 5 3% w/w gum arabicsoln. 15-20 — — — biocide 0.1 0.1 0.1 0.1 pH buffer (phosphate) 5-7 55-7 5 Phosphoric acid — 2 — 2 gum arabic EDTA — 2 — 2 EDTA — 0.5 — 0.5N. plumbaginifolia gum — — 0.3-0.4 — 3% (w/v) N. plumbaginofolia gum — —— 0.3-0.4

F1 and F2 are standard fountain solutions that employ gum arabic. F3 andF4 are identical to F1 and F2, respectively, except that N.plumbaginifolia gum product has been substituted for the gum arabic in aweight that is 1150 of the gum arabic weight.

When these fountain solutions were employed in an offset litho printing,it was found that the N. plumbaginifolia gum product performedcomparably to the gum arabic fountain solutions in terms of ink-plateroll up and in degree of plate background desensitization. The platewetting characteristics of the two products were also very similar. TheN. plumbaginifolia gum was found to be less soluble in isopropyl alcoholthan gum arabic; since isopropanol is very widely used as part of thedampening system of modern, fast lithographic offset presses, this maybe a negative feature.

Example 8 Fabric Printing—Preparation and Use of Reactive Dyestuff forWool or Cotton

Satisfactory dyeing of wool and cotton was accomplished as follows:

1. N. plumbaginifolia gum product 150 g 2. Cold water 2800 mL 3. Sodiummetaphosphate (Calgon ™) 30 g

The water was agitated with a high speed mixer during gradual additionof thesodium metaphosphate. The gum product was then added slowly, butfast enough so that all the powder was added before the viscosity hasrisen appreciably. Stirring was continued for 5-10 minutes until allparticles were swollen and had formed a thick suspension. The mixturewas allowed to stand for 1½ hours.

Then the following were added:

4. Diphasol ™ solution 115 mL 5. Hot water 975 mL 6. White spirit 3750mL 7. Resist salt L ™ 150 g

The thickening mixture was then stirred in the high speed mixer for 20minutes.

The screen printing paste was prepared by mixing the following:

1. Dyestuff 3 g 2. Urea 10 g 3. Hot to boiling water 30 mL 4. Thickening(as above) 50 g 5. Sodium bicarbonate 4 g

Using a high speed mixer, the dyestuff and urea were thoroughly drymixed. Then the hot water and thickening were added and mixed.

The printing paste was used in a standard fabric screen printing method.The printed cotton and wool were then dried followed by steaming for 8minutes. They were then rinsed thoroughly in cold water followed by asoaping at or near the boiling point with a detergent solution ofLissapol ND (2% w/w solution) and finally rinsed in cold water. Theprinting on the wool and cotton material appeared stable.

Example 9 Paints

9.A. Preparation of Acrylic Resin Paint

A stable water emulsion was prepared using the following formulations:

Premix in a ball mill:

1. Tap water 125 mL 2. Daxad 30 ™ Dispersant 8 g 3. Tergitol NPX ™Surfactant 4 mL 4. Victawet 35B ™ Wetting Agent 2.5 mL

Then the mill speed was increased and the following was added slowly:

5. Chemacoil TA-1001 ™ Resin 74 g

The speed was adjusted to disperse the following pigments and additives:

 6. Zinc oxide AZO-ZZZ-33 ™ 75 g  7. Titanox RANC ™ Rutile TitaniumDioxide 175 g  8. Titanox A168L0 ™ Anatase Titanium Dioxide 25 g  9.Asbestine 3X ™ Talc 100 g 10. Ethylene Glycol 18.5 g 11. Nuodex PMA-18Mildewoide ™ 3 g 12. Nopco NDW ™ Defoamer 4 g

The mill was then slowed to mixing speed.

13. N. plumbaginifolia gum product emulsion 165 mL (2% w/w aqueoussolution) 14. Rhoplex AC-34 ™ Acrylic Emulsion 372 g 15. Super Cobalt ™Drier 1 g

Mixing continued for at least ½ hour at mixing speed. Other pigments,such as carbon black or red oxide of iron, may be added to this toreplace part of the titanium dioxide ingredients in items 7 & 8 andprovide a differing color balance.

The above formulation was derived from Ernest Flick “Water-Based PaintFormulations” Noyes Publications, Parkridge, N.J.

9B. Oil Emulsion Paint

A satisfactory thixotropic paint was prepared as follows. Premix in ahigh speed stone mill:

1. Water 205 mL 2. Victawet 35B ™ Wetting agent 4 mL 3. Potassiumpolyphosphate 5 g

Adjust speed of the mill to disperse the following additives andpigments:

4. Ethylene glycol 10 g 5. Titanox RANC ™ Rutile TiO₂ 175 g 6. TitanoxA168L0 ™ Anatase TiO₂ 50 g 7. Asbestine 3X ™ Talc 50 g 8. Zinc oxideAZO-ZZZ-33 ™ 125 g 9. Nuodex PMA-18 Mildewoide ™ 2 mL 10.  Nopco NDW ™Defoamer 2 mL 11.  Victawet 35B ™ Wetting agent 16 g

The mill was then slowed to mixing speed and the following were added:

12. N. plumbaginifolia emulsion 130 mL (2.5% w/w) 13. Emulsified linseedoil 340 mL (60% solids) 14. Super Cobalt ™ drier 11 g

Milling was continued for approximately ½ hour.

This procedure resulted in a paint that tends to set to relatively stiffor “buttery” consistency upon standing, but thins down to relativelymobile liquid when mechanically agitated. This thixotropic character issuch that the shearing action of the brush used to apply the paint to asurface is sufficient to render the paint adequately mobile. The paintleaving the brush remains fluid for a sufficient time to bring aboutgood levelling (i.e., the brush marks disappear while the paint againsets to a stiff consistency before it has time to run appreciably on thesurface painted).

This thixotropic property in paints is valuable in flat paints meant tobe applied to interiors with a brush because it prevents the running ofthe paint and at the same time eliminates brush marks. Thixotropicpaints possess a further advantage quite apart from their intended usefor the reason above, in that the paint acquires a buttery or solidconsistency upon standing in containers. Segregation or stratificationof the paint during long periods of storage is thus prevented.

The above formulation and methods were derived from U.S. Pat. No.2,135,936, Nov. 8, 1938, for “Use of gum arabic in paint” and from“Emulsion and Water Soluble Paints and Coatings.”

Example 10 Ceramic Glazes

The suspension of the glaze ingredients in a glaze slip for severalhours or even days has been achieved using N. plumbaginifolia gumproduct as an emulsifying and/or suspending agent. Further, theresulting glaze has superior clarity and stability.

A stable glaze slip was prepared as follows:

To prepare an emulsifying/stabilizing mixture, the following werecombined:

1. N. plumbaginifolia dry gum product 10 g (grown on BLM as carbonsource) 2. Cold water 500 cc

Example 11 Clear Thixotropic Detergent or Cleaning Preparation

A satisfactory thixotropic cleaning detergent with superior grip andfilm fomring properties was prepared as follows:

1. Water 812 mL 2. N. plumbaginifolia (BLM carbon source) 40 g 3. Sodiumchloride 20 g

The gum product was added to the water in a high speed mixer running ata slow speed and was mixed for 15 minutes. The mixture was then left for1½ hours and the sodium chloride was added, mixing slowly for 3-5minutes. Sodium ethylsulphate was then added while mixing continued:

4. Sodium ethylsulphate 125 g (C12-14 2E.0.) (100% basis) 5. Perfume<0.5 g 6. Dye <0.5 g 7. Preservative <0.5 g

The perfume, dye and preservative were then added, and mixing continuedfor another 10 minutes.

Because the foregoing formulation does not contain either ethyl alcoholor propylene glycol (which can be used in cleaning detergents), thepossibility of precipitation of the gum due to a high alcoholconcentration is averted.

The resulting product is a clear cleaning agent which tends to berelatively stiff and provides an adequate detergent which clings to thesurfaces. This is a desirable characteristic particularly in thecleaning of vertical surfaces. It was found that the BLM carbon sourceenhanced the film forming properties of the detergent.

Example 12 Cosmetic Creams and Lotions

12.A. The concentrated gum from Nicotiana Batch 3-1000 (1.7% totalsolids) was found to have pleasant soft feeling on the skin and to drywithout stickiness. When mixed with water, it makes a satisfying, i.e.,moisturizing, skin treatment without any further additions. Anotherproduct was prepared by perfuming the biopolymer solution with 0.1% v/vrose oil.

12.B. A cosmetic lotion was prepared with the ingredients indicatedbelow. The vegetable oil, perfuming oil and glycerol were added to thebiopolymer solution while mixing with a high speed stirrer such as anUltraturrax at a setting of about 6.

Nicotiana gum #3-1000 2.4% (w/w in H₂O) Orange oil 1.0% (w/w) Olive oil2.3% (w/w) Glycerol 5.3% (w/w)

The Nicotiana gum was mixed in the water in a high speed stirrer such asan Ultraturrax at a setting of about 6. The olive oil, orange oil andglycerol were then added. The result was a soft gel with a pleasantfresh aroma which can be spread on the hands or face, leaving skinfeeling fresh and soft.

12.C. A cosmetic lotion was prepared using the following:

Nicotiana gum #3-1000 1.7% (w/w in H₂O) Sunflower oil 1.3% Glycerol 4.0%

The ingredients were combined as in 11.B. The resulting product was softenough to be used in a pump-action dispenser. A perfuming oil can beadded if desired.

12.D. Other batches of gum from other cell lines were used to prepareproducts with different properties. For example, a cream was preparedfrom gum produced by Nicotiana cells growing in a medium containingBrewers Liquid Maltose (“BLM”) 162 g/liter, as the source of sugar. Theresulting gum produced a viscous solution and was used to prepare alotion with the following formulation:

Nicotiana gum 2.3% (w/w in H₂O) Peanut oil 3.3% Rose oil 0.1% Glycerol5.0%

Example 13 Compositions and Selected Rheological Properties of PlantCell Gums

Table 4 lists relative weight proportions of protein and ash and typesof polysaccharide in plant cell gums obtained from suspension culturesof a variety of vascular plants including both dicots and monocots.Table 5 lists explant sources and culturing conditions forplant cellcultures of Tables 4 and Table 6 lists the maximum polysaccharideconcentration obtained in cultures during growth cycle measurements.

There is significant variation in the amount of protein+ash, from a lowof about 12% for Timothy Grass to a high of 58% for white clover, in therepresentative plant cell gums exemplified. Xyloglucan is relatively lowin the gums of cells of Fabaceae and Poacea but relatively high inAizoaceae and Malvaceae plant cell gums. Arabinogalactan which will beattached to protein also varies significantly from a high of 28% to alow of 10%. Heteroxylan is only significant in cells of monocots.Glucuronomannan is high in the monocot Poacea, but also in Nicotianaplumbaginifolia plant cell gum. Rhamnogalacturonan is also variableamong the gums assessed.

The percentages by weight of the components measured add upapproximately to 100% for each cell line. Discrepancies in total weightpercent may occur due to the inherent errors in analytical techniques orif the assumptions made with respect to the structures ofpolysaccharides present are inaccurate. For example, the presence ofminor functional groups such as methyl or acetyl groups may not be takeninto account. The relative amounts of individual polysaccharides isinferred from detailed methylation data from isolated gum.

Methylation analysis was performed using analytical methods well-knownin the art. Protein and ash were also measured using standard methodswell-known in the art. Briefly, total nitrogen (N) was determined usingthe Kjeldahl method and expressed as protein (N×6.25). Inorganicmaterial (ash) was determined by ash content. Monosaccharidecompositions were determined by GC/MS following carboxyl reduction andmethylation analysis. Isolated gums were carboxyl reduced and methylatedand the partially methylated alditol acetates were analyzed by GC-MS.

The proportions of individual polysaccharides present in the gum weredetermined based on characteristic linkage structures of purifiedpolymers from dicotyledonous and monocotyledonous species as describedby Shea et al. (1989) Planta 179:293. Xyloglucan was the sum of 4,6-Glcand terminal Xyl equal to 4,6-Glc, as well as 2-Xyl, 2-Gal, terminal Fucand 4-Glc equal to one third that of 4,6-Glc, except for Solanumtuberosum and Lycopersicon esculentum in which the structure ofxyloglucan from S. tubersum was used as a model (Ring and Selvendran(1981)). Galactoglucomannan was the sum of 4-Man and 4,6-Man, 4-Glcequal to the sum of these Man linkages and terminal Gal equal to4,6-Man. Glucomannan was the sum of 4-Man and any 4-Glc not assigned toxyloglucan. 3,6-Arabinogalactans (type II) were the sum of 3-Gal, 6-Gal,and 3,6-Gal and terminal Ara equal to 3,6-Gal. Heteroxylans were the sumof 4-Xyl, 2,4-Xyl,and 3,4-Xyl and terminal GlcA and terminal Ara equalto 2,4-Xyl and 3,4-Xyl. 4-Galactan was the sum of 4-Gal and 2,4-Gal andterminal Gal equal to 2,4-Gal, Arabinan was the sum of 2-Ara, 3-Ara,5-Ara, 2,5-Ara and terminal Ara equal to 2,5-Ara. Glucuronomannan wasthe sum of 2-Man, 2,3-Man and 4-GlcA, 3,4-GlcA and terminal Ara andterminal Gal equal to the sum of 2,3-Man and 3,4-GlcA. Galacturonan wasthe sum of 4-GalA, 4-GalA(6-O-Me) and 3,4-GalA, and rhamnogalacturonanwas the sum of these linkages and 2,4-Rha.

Suspension cultures were initiated from explants indicated in Table 6generally following the methods described in Example 2 except thatdifferent media, hormone balance (as indicated in Tables 5 and 6) andfermentation times were used. Suspension cultures were grown in 2L shakeflasks shaken at 100 rpm at 27° C. in the dark.

Table 6 lists gum yield produced by suspension cultures. Gum productionin these cell lines has not as yet been optimized by variation offermentation conditions and media components. Increased yields of gumcan be achieved by such optimization methods.

Table 4 lists emulsification, gelling and the viscosity of various dicotand monocot plant cell gums. Gums were isolated essentially as describedin example 2. Emulsification was tested by determining the droplet sizeof a limonene emulsion formed using 1% (w/v) of an aqueous gum solutionessentially as described in example 3. Mesembryanthemum gum gave anexcellent emulsion with low droplet size of 4.8 μm which is three tofive times smaller than emulsion droplets formed using Nicotiana andPyrus gum, respectively. Gums which can generate an emulsion withdroplet size less than about 5.0 μm are particularly useful in the drinkindustry. Low-droplet-size emulsions are those in which droplet size is5.0 μm or less; preferred low-droplet-size emulsions are those havingdroplet size less than about 2.0 μm.

Viscosity was measured essentially as described in example 3B. Some ofthe gums including timothy grass and millet appear to be viscoelastic,i.e., displaying both solid-like (elastic) and liquid-like (viscous)properties. Viscoelastic properties are time dependent. A viscoelasticmaterial can exhibit either linear or non-linear viscoelastic behavior.Viscoelastic behavior can be observed, for example, by rapidly twistinga bottle of gum and watching recoil. Linear viscoelastic behavior isobserved at low strain (non-destructive testing). Measurements ofstorage modulus(G′) and loss modulus (G″), both well-known measurements,give an indication of the amount of solid-like behavior (G′) and liquidbehavior (G″).

Mesembryantheum gum has a relatively low viscosity combined with thecapability to form low droplet size emulsions. Mesembryantheum gum isthus useful in emulsification applications and particularly well-suitedfor use in cloud emulsions. Cloud emulsions have applications in thefood industry, for example for manufacture of soft drinks, and in thechemical industry and agriculture for preparation of chemical emulsionsincluding emulsions of agricultural chemicals (pesticides, etc.). Gumsof species of Aptenia and Carpobrotus are also well suited forpreparation of emulsions.

Timothy grass gum displays good gelling ability and high viscosity. Thisgum is useful in applications where there is a need to suspend orstabilize or where water-holding it properties are needed. This gum hasapplication, among many others, in the food industry for ice cream anddessert items and in the preparation of spray emulsions, for example,for agricultural chemicals.

Millet gum has high viscosity associated with water thickeningproperties. In addition to a variety of uses in the food industry insauces, bakery glaces and other foods, millet gum has application in thepreparation of drilling muds.

Derek Morton “Determination of Primary Structure of MicrobialPolysaccharides”, Ohio Food Hyrocolloids Conference, September 1994,provides an exemplary listing of applications and specific industrialuses for gum having a given rheological property or characteristic.

Example 14 Additional Media for Plant Cell Cultures

Linsmaier and Skoog Powder (LS) Medium for Chicory

LS Powder 4.16 g Sucrose 30.0 g Organics stock 1.0 mL 2,4-D (0.1 mg/mL)2.2 mL NAA (0.2 mg/ml) 0.186 mL

Dissolve all ingredients, except agar, in 800 mL Milli-Q water. AdjustpH to 6.0 with KOH and make up to 1 L. Add agar 5.0 g if making solidmedium. Autoclave for 20 m at 121° C.

Organic Stock Thiamine Hydrochloride 40 mg Myo-inositol 10 g

Dissolve both ingredients and make up to 100 mL with Milli-Q water anduse stock at 1 mL/L. Aliquot into 10 mL portions and freeze.

Tomato BAL Medium

Add concentrated stock solutions and sucrose to approximately 900 mL ofMilli-Q water with constant agitation using a magnetic stirrer:

Macro elements 10 mL Micro elements 1 mL Vitamins 1 mL CaCl₂ 2.6 mLKH₂PO₄ 10 mL Fe.EDTA 1 mL 2iP 1 mL 2,4-D 20 mL KI 0.5 mL Sucrose 30 g

Macro and micro elements and vitamins are the same as used in pear BAL.Adjust pH to 5.0-6.0 with KOH (0.1 or 1M) and adjust final volume to 1liter. For solid medium add 0.5% agar (5 g/L) after adjusting Ph andvolume. Sterilize for 20 m at 10 psi (116° C.).

TB1 Medium for Tuberose (PETAL)

LS powder 4.60 g Sucrose 30.0 g 2,4-D 20.0 mL (Stock Solution 0.1 mg/L)

Dissolve all ingredients in approximately 800 mL Milli-Q water and thenmake up to 1 L. Adjust pH to 5.8-6.0 with 0.1M KOH. To prepare solidmedium, add 5.0 g of agar to a 2 L flask, add dissolved consitutents andsteam until dissolved. Sterilize at 10 psi for 20 m.

A related medium TB2 (Tuberose 2) also used for petal culture isprepared in a similar manner but contains {fraction (1/10)}th the amountof 2,4-D (i.e., add 2.0 mL of the Stock Solution of 2,4-D rather than 20mL).

Example 15 Production of Plant Cell Gum of Mesembryanthemum “Pigface”

A commerically available succulent plant with orange flowers designatedMesembryanthemum purchased in 1992 from Sherringham's Nursery (New Ryde,NSW, Australia) as supplied by Milingimbi Nursery (Jillibi, NSW,Australia) was employed to establish a Mesembryanthemum suspension cellculture. This plant is commonly called pigface and is a very commonlandscaping plant in the Central Coast region of New South Wales,Australia. It is possible that this plant is a cross betweenMesembryanthemum and/or Carpobrotus or that it is more correctlydesignated Carpobrotus. The labelling employed by the nursery where theplant was purchase is used herein.

Callus was initiated from leaf tissue cultured on Murashige's and Skoogmedium (MS2) containing 2.0 mg/L 2,4-D ( see Murashige.T and Skoog, P.(1962) Physiol. Plant 15 473-97) medium solidified with 0.5% (w/w) agar.Subculturing was performed substantially as described in Example 1wherein the number of subcultures of callus is indicated in Table 5.Suspension cultures are established employing portions of callus tissuewherein the number of subcultures are indicated in Table 5.

MS2 or a modified MS2 medium having double the concentrations of allorganics, phosphate and all trace elements designated PF2 have been usedfor suspension cultures of Mesembryanthemum. Suspension cell media aretypically provided with 2.0 mg/L the plant hormome 2,4-D. Cells aretypically cultured at temperatures ranging from about 25-28° C. withdissolved oxygen maintained at 20-70% air saturation. Culture pH istypically not controlled externally.

MS2 PF2 Main Inorganics (g/L) (g/L) NH₄NO₃ 1.65 1.65 KNO₃ 1.9 1.9 CaCl₂· 2H₂O 0.44 0.44 MgSO₄ · 7H₂O 0.37 0.37 KH₂PO₄ 0.17 0.34 Trace Elements(mg/L) (mg/L) KI 0.83 1.66 H₃BO₃ 6.2 12.4 ZnSO₄ · 7H₂O 8.6 17.2 MnSO₄ ·4H₂O 22.3 44.6 Na₂MoO₄ · H₂O 0.25 0.5 CuSO₄ · 5H₂O 0.025 0.05 CoCl₂ ·6H₂O 0.025 0.05 Iron Source (mg/L) (mg/L) FeSO₄ · 7H₂O 27.8 27.8 Na₂EDTA· 2H₂O 37.3 37.3 Organic Supplements (mg/L) (mg/L) Myo-Inositol 100 200Nicotinic acid 0.5 1 Pyridoxiene-HCl 0.5 1 Thiamine-HCl 0.1 0.2 Glycine2 4

Plant cell gum was isolated from Mesembryanthemum suspension culturessubstantially as described in Example 2. Briefly, cells (biomass) areseparated from culture filtrate, and Na₂EDTA.2H₂O (1 g/L) (as asequestering agent) is added to the filtrate and the pH is adjusted toabout 8 to complex calcium. Preservatives and antioxidants (ascorbicacid, potassium sorbate, sodium benzoatesodium metabisulfate, etc.) canbe added to the culture filtrate to prevent deterioration of gumcomponents. Treated filtrate is concentrated using ultrafiltration or ina dialysis sack covered with polyethylene glycol flakes to ⅓ or ¼ itsoriginal volume. Concentrated filtrate is washed (by diafiltration or bydialysis). The washed filtrate is dried for example by spray drying orfreeze drying.

Dried gum can be assessed for emulsification capacity and inert fillers(such as maltodextrin) can be added to dried gum to provide astandardized sample having a selected emulsification power.

Mesembryanthemum plant cell gum for use in food or veterinaryapplications can be further purified by alcohol precipitation if desiredor necessary.

FIG. 1 is a plot comparing emulsification capacity of Mesembryanthemumgum with that of a commercial spray-dried gum arabic. Emulsion dropletsize (DWO) in gm is plotted as a function of gum concentration.Mesembryanthemum gum (squares) gives a significantly betteremulsification, i.e., small droplet size, compared to gum arabic(circles). Mesembryanthemum gum also exhibits good emulsification atrelatively low concentration (approaching 0.1% (by weight).

Example 16 Splash Inhibition Studies of Millet Plant Cell Gum

The formation of fine mists and satellite droplets is undesirable incertain agricultural chemical compositions, such as those containingpesticides, herbicides and fungicides. Formation of fine mists onchemical application to fields or gardens can result in undesired widedispersion of hazardous materials. The use of components such aspolyethylene oxide in agricultural chemical compositions can inhibitformation fine mists and satellite droplets. Satellite dropletinhibition is also important in ink applications, for example, inapplication to ink-jet printing. Monocot plant cell gums, particularlythose of millet and timothy grass are found to have viscoelasticproperties which are associated with fine mist inhibition and satellitedroplet inhibition. Splash inhibition assays can be used to assess thepotential of a plant cell gum for use in applications needing fine mistand satellite droplet inhibition.

Splash inhibition in solutions containing millet plant cell gum (as inTable 4) was examined in a drop impact study in which millet solutionswere compared to solutions containing polyethylene oxide (PEO). A 35%aqueous glycrol solution containing 0.15% millet plant cell gum wasfound to have a shear viscosity of about 5.5 cP at 25° C. Aqueousglycerol (35%) solutions containing PEO (either 8,000 Mwt, 300,000 Mwt,600,000 Mwt, or 1,000,000 Mwt PEO with concentration [0.1%-0.17%]dependent upon molecular weight of the PEO) having the same shearviscosity (5.5 cP at 25° C.) were compared to the millet solution. A 50%aqueous glycerol solution was also tested. Glycerol was included insolutions to enhance viscosity for measurement purposes. The Troutonratio of the millet solution at 25° C. is about 120 compared to that ofa 1,000,000 Mwt. PEO solution which is 5.5. The millet gum solution isabout 20-fold more elastic than the PEO solutions.

All of the solutions examined for splash inhibition have approximatelythe same shear viscosity, density, surface tension drop diameter andimpact velocity. The solutions differ in extensional viscosity. At highextensional rates the extensional viscosity of the millet solutiondecreases compared to PEO solutions.

Solution drops are impacted on aluminum surfaces roughened with varyinggrades of sandpaper (500 grit to 40 grit). The initial drop onto thesurface does not splash; splashing is observed when a drop hits a thinfilm of liquid, e.g., when the second drop impacts the wet surface. Theimpact of a drop is captured by a progressive scan video camera at 50frames/sec.

The 50% aqueous glycerol solution and the 8,000 Mwt. PEO (in 35% aqueousglycerol) solution splashed on all aluminum surfaces tested at both 20and In contrast, solutions containing higher molecular weight PEO(300,000-1,000,000 Mwt) do not splash on the tested surfaces at either20 or 25° C. The millet solution (0.15% millet plant cell gum in 35%aqueous glycerol) does not exhibit splashing at 20° C., but does exhibitsplashing at 25° C. The millet plant cell gum solution tested doesexhibit splash inhibition and can be employed in compositions to provideinhibition of fine mists and satellite droplets.

Those of ordinary skill in the art will appreciate that materials mediacomponents, purification methods, techniques and procedures other thanthose specifically exemplified herein can be employed in the practice ofthis invention without departing from its spirit and scope which isdefined in the appended claims.

TABLE 1A Exemplary Plant Sources for Cultured Plant Cell Gums CLASS:MAGNOLIOPSIDA (DICOTS) SUBCLASS 1: MAGNOLIDIAE ORDER 1: MAGNOLIALESFamily: Annonaceae Annona muricata (custard apple) Family: MagnoliaceaeMagnolia grandiflora ORDER 2: LAURALES Family: Lauraceae (Cassythaceae)Persea americana (avocado) Cassytha melantha, Aust native Cassythapubescens, Aust native Cassytha glabella, Aust native ORDER 3: PIPERALESFamily: Piperaceae Pepperomia obtusifolia ORDER 4: ARISTOLOCHIALESFamily: Aristolochiaceae Tropical climbers ORDER 5: ILLICIALES Family:Schisandraceae Schisandra spp., climbers Kadsura spp. ORDER 6:NYMPHAEALES Family: Nymphaeceae Nymphaea spp., water lilies Braseniaschreberi, water plant Family: Nelumbonaceae Nelumbo spp., lotus ORDER7: RANUNCULALES Family: Ranunculaceae Ranuncula bulbs Family:Lardizabilaceae Akebia quinata, climber Family: Berberidaceae Berberisspp., shrub Mahonia spp., huckleberry ORDER 8: PAPAVERALES Family:Papaveraceae Papaver somniferum, poppy Family: Fumariaceae Fumaria spp.,weed herb SUBCLASS 2: HAMAMELIDAE ORDER 2: HAMAMELIDALES Family:Hamamelidaceae Liquidambar Hamamelis spp., witch hazel Family:Plananaceae Planatus spp., plane tree ORDER 6: URTICALES Family:Cannabaceae Cannabis sativa, hemp Family: Urticaceae Boehmeria nivea,ramie ORDER 8: JUGLANDALES Family: Juglandaceae Juglans spp., walnutCarya spp., pecan ORDER 9: MYRICALES Family: Myricaceae Comtoniaperegrina, ornamental ORDER 10: FAGALES Family: Betulaceae Betavulgaris, sugar beet Beta vulgaris, beetroot Betula verrucosa, birchFamily: Nothofagaceae Nothofagus moorei, Aust. beech Family: FagaceaeFagus sylvatica, Europ. beech ORDER 11: CASUARINALES Family:Casuarinaceae Casuarina spp. SUBCLASS 3: CARYOPHYLLIDAE ORDER 1:CARYOPHYLLALES Family: Aizoaceae Mesembryanthemum chilense (original) 4presumed (ID undertaken) varieties Aptenia cordii Carpobrotusacinaciformis Carpobrotus edulis Delosperma lehmanni Hereroa dyeriRushia rubricaulis Family: Chenopodiaceae Spinacia oleracea, spinachFamily: Basellaceae Basella alba, San Choy Family: CactaceaeEchinocactus grusonii Neoporteria cormasensis Opuntia dillenii, pricklypear Family: Amaranthacea Amaranthus spp. Family: CaryophyllaceaeDianthus caryophyllus ORDER 2: POLYGONALES Family: PolygonaceaePolygonum spp., weed Persicaria spp., weed ORDER 3: PLUMBAGINALESFamily: Plumbaginaceae Plumbago spp., shrub Limonium spp., cut flowerSUBCLASS 4: DELLENIIDAE ORDER 1: DILLENIALES Family: Paeoniaceae Paeonyspp., cut flower ORDER 2: THEALES Family: Theaceae Camellia japonicaFamily: Actinidiaceae Actinidia chinensis, kiwi fruit Family: ClusiaceaeHypericum perforatum, St. John's Wort Hypercium androsaemum, tutsanORDER 3: MALVALES Family: Tiliaceae Corchorus spp., jute Tilia spp.,ornamental Family: Malvaceae Gossypium arboreum, cotton Gossypiumhirsutum, cotton Hibiscus cannabinus, mesta Hibiscus sabdariffa, rozellaHibiscus esculentus, okra Sida rhambifolia, Paddys' lucern Alathaeaspp., marshmallow, bioemuls Abelmoschus glutinotextilis Family:Sterculiaceae Sterculia urens, Karaya gum ORDER 4: LECYTHIDALES Family:Barringtoniaceae Barringtonia spp., Aust. native Family: LecythidaceaeBerthollethia spp., Brasil nut ORDER 5: NEPENTHALES Family:Sarraceniaceae Sarracenia spp., Pitcher plant Family: DroseraceaeDrosers spp., native carnivorous ORDER 6: VIOLALES Family:Passifloraceae Passiflora edulis, passion fruit Family: CucurbitaceaeSechium edule, choko Cucumis sativus, cucumber Crystal Apple Cucumissativus, cucumber Burpless Cucurbita pepa, zucchini Cucurbita maxima,butternut pumpkin Citrullus lanatus, watermelon Family: Violaceae Viiolaodorata, violet Family: Begoniaceae Begonia spp., begonia ORDER 7:SALICALES Family: Salicaceae Populus tremuloides, aspen ORDER 8:CAPPARALES Family: Brassiaceae Brassica hirta, yellow mustard Brassicaoleracea, cabbage Brassica sinapsis, white mustard Sinapsis alba,mustard Arabidopsis thaliana Family: Mocingaceae Moringa petrygosperma,horseradish tree ORDER 9: BATALES Family: Gyrostemonaceae Gyrostemonspp., Aust. natives ORDER 10: ERICALES Family: Ericaceae HeathRhododendron Vaccinium myrtillus, bilberry Family: Epacridaceae Epacrisspp., Vic native Leucopogon spp., Vic native ORDER 12: EBANALES Family:Ebenaceae Diospyros virginiana, persimmon Family: Sapotaceae Palaquimspp. guttapercha Payena spp., guttapercha Chrysophyllum spp., chewinggum Manilkara spp., chewing gum ORDER 13: PRIMULALES Family: PrimulaceaeCyclamen euroeum Primula SUBCLASS 5: ROSIDAE ORDER 1: ROSALES Family:Rosaceae Malus pumila, apple Malus domesticus cv Braebum Rosa glaucaPrunus avium, sweet cherry Prunus insitia, damson Prunus domestica, eggplum Prunus cerasus, cherry Prunus virginiana, cherry Prunus persica,peach Prunus amygdalus, almond Prunus armenica, apricot Pyrus communisFragaria ananassa, strawberry Vaccinium macrocarpon, cranberry Family:Cunoniaceae (Baueraceae) Bauera spp., Aust native Family: PittosporaceaePittosporum undulatum Billardiera scandens, Aust. native Sollyaheterophylla Family: Hydrangeaceae Hydranges spp. Family:Grossulariaceae Ribes uva crispa, gooseberries Ribes rubrum, currantsRibes nigrum, blackcurrants ORDER 2: FABALES OR LEGUMINOSAE Family:Fabaceae Anthyllis vulneraria, kidney vetch Astragalus cicer Astragalusglycyphyllos Astragalus gummifera, tragacanth Astragalus nuttallianusAstragalus sinicus Astragalus tenellus Centrosoma plumari Certoniasiliqua, carob Cercidium torreyanum, palo verde Crotalaria incanaCrotalaria intermedia Crotalaria juncea Crotalaria lanceolata Crotalariamedicaginea Crotalaria mucronato Crotalaria retusa Crotalariaspectabilis Crotalaria striata Cyamopsis tetragonoloba, gar Delonixregia Demanthus pulchellum Desmodium pulchellum Genista raetam Genistascoparia Glycine max, soy bean Gymnocladus dioica, Kentucky coffeeIndigofera hirsutum, Indian legume Leucaena glauca, Koa hoale Lotuscorniculatus Lotus peduculatus Lotus scoparius Lupinus albus Lupinusangustifolius Lupinus luteus Medicago hispida Medicago Lupulina Medicagoorbicularis Medicago radiata Medicago sativa, alfalfa Melilotus albusMelilotus indica Mimosa scabrella Mucuna imbricata Parkinsonia aculeataPhaseolus aureus, mung bean Phaseolus atropurpureus, Sirato Phaseolusvulgaris Sesbania grandiflora Sesbania speciosa Sesbania bispinosaSophora japonica Stylosanthes humilis, Townsville lucerne Tamarindusindica, tamarind Tetragonolobus purpurens Trifolium alexandrinumTrifolium dubium Trifolium hirtum Trifolium incarnatum Trifolium repens,white clover Trifolium resupinatum Trifolium pratense, red cloverTrigonella caerulea Trigonella cretica Trigonella calliceras Trigonellacorniculata Trigonella foenum-graecum, fenugreek Trigonella hamosaTrigonella monspeliaca Trigonella polycerata Vicia faba Vicia sativaProsopis velutina, mesquite Alysicarpus vaginalis Virgilia oroboides,trees exuding gum Virgilia divaricata Family: CaesalpiniaceaeCaesalpinia cacalaco Caesalpinia pulcherima C. spinosa, Tara Gleditsiaamorhoides G. Ferox G. triacantos Cassia emarginata C. fistula C. absusC. leptocarpa C. marylandica C. nodosa C. occidentalis C. tora Family:Mimosaceae Mimosa scabrella Acacia dealbata Acacia mearnsii Acacianilotica Acacia pycnantha Acacia senegal Acacia saligna Albizialophantha Albizia julibrissin ORDER 3: PROTEALES Family: ProteaceaBanksia integrifolia, gooey fruits Grevillea, gooey fruits Hakea spp.,gooey fruits Dryandra spp., gooey fruits Macadamia spp., gooey fruitsPersoonia spp., gooey fruits ORDER 5: HALORAGALES Family: HaloragaceaeMyriophyllum spp., aquarium plant ORDER 6: MYRTALES Family: OnagraceaeFuchsia spp., fuchsia Oenothera biennis, evening primrose Family:Myrtaceae Eucalyptus camaldulensis Eucalyptus gunni Leptospermum spp.Thriptomene spp. Eugenia spp., Lillipilli Callistemon spp. Kunzea spp.Mellaleuca spp. Family: Punicaceae Punica granatum, pomegranite Family:Combretaceae Combretum collinum Combretum hartmanniana Combretumleonense Anogeissus latifolia, gum ghatti Anogeissus leicocargus ORDER7: RHIZOPHORALES Family: Rhizphoraceae Rhizophora spp., mangrove ORDER8: CORNALES Family: Cornaceae Cornus spp., dogwood Davidia involucrataFamily: Garryaceae Garrya ellipitica ORDER 9: SANTALES Family:Santalaceae Exocarpus spp., Aust native cherry Family: LoranthaceaeMistletoe amyema Family: Viscaceae Mistletoe dendrophora Korthalsellaspp. ORDER 11: CELASTRALES Family: Aquafoliaceae Ilex aquifolium, hollyFamily: Stackhousiaceae Stackhousia monogyna, Aust native ORDER 12:EUPHORBIALES Family: Buxaceae Buxus spp., European box Family:Euphorbiaceae Euphorbia spp., native weed ORDER 13: RHAMNALES Family:Rhamnaceae Rhamnus spp., black juicy fruit Family: Vitaceae Vitisvinzftra, grapes ORDER 14: LINALES Family: Linaceae Linum usitatissimum,flax ORDER 15: POLYGALALES Family: Polygalaceae Polygala spp. Family:Tremandaceae Tetratheca spp., Aust shrub ORDER 16: SAPINDALES Family:Aceraceae Acer pseudoplatinus Acer saccharum Family: Meliaceae Khayasenegalensis, Khaya gum Family: Anacardiaceae Schinus molle, peppercorntree Mangifera spp., mango Anacardium spp., cashews Pistachia spp.,pistachio ORDER 17: GERANIALES Family: Tropaeolaceae Tropaeolum majus,nasturtium Family: Balsaminaceae Impatiens balsamina Family: OxalidaceaeOxalis spp., weed Family: Geraniaceae Geranium spp. Pelargonium spp.ORDER 18: APIALES Family: Apiaceae Carum carvi, carroway Daucus carota,carrot SUBCLASS 6: ASTERIDAE ORDER 1: GENTIANALES Family: AsclepiadaceaeAraujia hortum Family: Apocynaceae Vinca major, blue periwinkle ORDER 2:SOLANALES Family: Solanaceae Nicotiana alata Nicotiana sylvestrisNicotiana tabacum Lycopersicon esculentum, tomato Solanum tuberosum,potato Family: Convolvulaceae Convolvulus tricolor Ipomoea muricataIpomoea batatas, sweet potato ORDER 3: LAMIALES Family: BoraginaceaeMyosotis spp., Forget-me-not Echium vulgare, Paterson's curse Boragooffinalis, borage Symphytum officinale, comfrey Family: Lamiaceae Mesonaprocumbens, Hsian tsao Prosanthera spp., Aust native Westringia spp.,Aust native Lavandula spp., lavender Mentha spp., mint ORDER 5:PLANTAGINALES Family: Plantaginaceae Plantago indica Plantago afra,psyllium ORDER 6: SCROPHULARIALES Family: Buddlejaceae Buddleja spp.,butterfly bush Family: Acanthaceae Acanthus spp. Family: MyoporaceaeMyoporum spp., Aust native Family: Scrophulariaceae Antirrhinum spp.,snapdragons Digitalis spp., foxgloves ORDER 7: CAMPANULALES Family:Goodeniaceae Goodenia spp., Aust native Dampiera spp., Aust nativeFamily: Campanulaceae Wahlenbergia spp., bluebells ORDER 8: RUBIALESFamily: Rubiaceae Coffea arabica, coffee Coprosma spp., Aust nativeGardenia spp ORDER 9: DIPSACALES Family: Caprifoliaceae Lonicera spp.,honeysuckle Viburnum opulus Abelia spp. ORDER 11: ASTERALES Family:Asteraceae Lactuca sativa, lettuce Helianthes annus sunflower Taraxacumofficinale, dandelion Cichorium intybus, chicory Cynara scolymus,Jerusalem artichoke Silybum marianum, silymarin Echinacea augustofoliaOther dicot species of uncertain classification Simmondsia chinensis,jojoba Moraceae Artocatpus heterophyllus, jackfruit NyctaginaceaePisonia spp., para para CLASS: LILIOPSIDA (MONOCOTS) SUBCLASS 1:ALISMATIDAE ORDER 3: MAJADALES Family: Zosteraceae Phyllospadix spp.,sea grass Zostera marina Zostera annuus SUBCLASS 2: ARECIDAE ORDER 1:ARECALES Family: Arecaceae (Palmae) Arenga saccharifera Borassusflabellifer Cocos nucifera, coconut Phytelephas macrocarpa, Ivory nutElaeis guineensis, oil palm Hyphaene thebaica, doum palm Phoenixdactyliflora, date palm ORDER 4: ARALES Family: Araceae Monsteradeliciosa, monstera Amorphophallus konjac SUBCLASS 3: COMMELINIDAE ORDER1: COMMELINALES Family: Commelinaceae Tradescantia albiflora, wanderingjew SUBCLASS 4: ZINGERBERIDAE ORDER 1: BROMELIACAE Family: BromeliaceaeAnanas comosus, pineapple SUBCLASS 3: COMMELINIDAE ORDER 5: CYPERALESFamily: Poaceae (Graminaceae) Avena sativa, oats Hordeum vulgare, barleyLolium multiflorum, rye grass L. perenne, rye grass L. temulentum, ryegass Oryza sariva, rice Panicum miliaceum, millet Saccharum officinarum,sugar cane Triticum aestivum, wheat Zea mays, corn Phleum pratense,Timothy grass Secale cereale, rye SUBCLASS 5: LILIDAE ORDER 1: LILIALESFamily: Liliaceae Allium cepa, onion Allium sativum, garlic Asparagusofficinalis, asparagus Tulipa gesneriana, tulip Edymiom nutans, nurserybulb Lilium longiflorum Scilla nonscripta, nursery bulb Aloe veraHomesia miniata, cape tulip Family: Iridaceae Iris ochroleuca, iris Irissibirica Watsonia pyrimidata Gladiolus spp. Family: Agaveaceae Agavesapp., succulent Agave sisalana, sisal Cordyline indivisa Sansevierratrifasciata Polyanthes tuberosa, tuberose Family: HaemodoraceaeAnigozanthus spp., kangaroo paw CLASS: GYMNOSPERMS ORDER: CONIFERALESFamily: Pinaceae Abies balsamea, Balsam fir Picea abies, spruce Piceasitchenis, Sikta spruce Pinus resinosa, red pine Pinus sylvestris, pineLarix occidentalis, larch Pseudotsuga menziesii Family: CupressaceaeCupressus spp. Family: Podocarpaceae Podocarpus elatus, plum pine ORDER:GINKOGOALES Ginkgo biloba Species of uncertain classification Yew Inulahelenium, elecampane Pouteria cainito, abiu

TABLE 1B Preferred Plant Cells for Cultured Plant Cell Gum ProductionFamilies Actinidiaceae Agavaceae Aizoaceae Asteraceae BoraginaceaeBrassicaceae Caesalpiniaceae Convolvulaceae Cucurbitaceae FabaceaeGeraniaceae Malvaceae Mimosaceae Myrtaceae Passifloraceae Palmae PoaceaePrimulaceae Rosaceae Solanaceae Tropaeolaceae Genera Acacia ActinidiaAptenia Arabidopsis Avena Cactaceae Caesalpinia Carpobrutus CichoriumCocos Cucumis Cucurbita Cynara Echinacea Echinocactus Eucalyptus GlycineHibiscus Hordeum Ipomoea Lactuca Lycopersicon Malus MedicagoMesembryanthemum Mimosa Neoporteria Nicotiana Oryza Panicum PassifloraPelargonium Phelum Polyanthes Primula Pyrus Rosa Sida Solanum SymphytumTaraxacum Trifolium Trigonella Triticum Tropaeolum Vaccinium Zea SpeciesAcacia dealbata Acacia senegal Actinidia chinensis (kiwi fruit) Apentiacordifolia Aptenia cordii Arabidopsis thaliana Avena sativa Caesalpiniapulcherima Carpobrotus acinaciformis Carpobrotus aequilaterusCarpobrotus chilense (also Mesembryanthemum chilense) Carpobrotusconcavus Carpobrotus deliciosus Carpobrotus dimidiatus Carpobrotusdisparilis Carpobrotus dulis Carpobrotus edulis Carpobrotus fourcadeiCarpobrotus glaucescens Carpobrotus laevigatus Carpobrotus melleiCarpobrotus modestus Carpobrotus muirii Carpobrotus pageae Carpobrotuspillansii Carpobrotus pulleinii Carpobrotus quadrifidus Carpobrotusrossii Carpobrotus rubrocinctus Carpobrotus sauerae Carpobrotus sublatusCarpobrotus vanzijlae Carpobrotus virescens Carpobrotus spp. Cichoriumintybus Cocos nucifera Cocos nucifera (makapuno mutant) Cucumis sativusCucurbita maxima Cynara scolymus Echinacea augustafolia Echinocactusgrusonii Eucalyptus camaldulensis Glycine max Hibiscus esculentusHordeum vulgare Ipomoea batatas (sweet potato) Lactuca sativaLycopersicon esculentum Malus domesticus Medicago sativaMesembryanthemum aitonis M. alatum M. albatum M. alborroseum M. annum M.barklyi M. breve M. chrysum M. clandestinum M. cordifolium M.criniflorum M. cryocalyx M. cryptanthum M. crystallinum M. dejagerae M.edulis (Carpobrotus edulis) M. excavatum M. forsskalei Hochst M.galipinii M. guerichinanum M. hypertrophicum M. inachabense M. inornatumM. intransparens M. karrooense M. latisepalum M. liebendalense M.lincarifolium M. longipapillosum M. louiseae M. macrophyllum M.macrostigma M. nellsonlae M. nodiflorum M. pachypus M. parvipapillatumM. paucandrum M. paulum M. pellitum M. perlatum M. purpureo-roseum M.quinangulatum M. rhodanthum M. rubroroseum M. sedentiflorum M. setosumM. squamulosum M. stenandrum M. subrigidum

TABLE 2 Droplet size and turbidity measurement Conc Droplet Turbidity (&w/v) size (μm) ABS 500 nm Abs-Avr Gum arabic  0 A Very large 0.049,0.023 0.002 B Very large 0.013, 0.003  0.2 A Very large 0.006, 0.0090.023 B 10-20 0.058, 0.022  0.5 A 10-20 0.042, 0.040 0.038 B 4-20 0.034,0.037  1 A 6-20 0.145, 0.130 0.128 B 3-12 0.125, 0.112  5 A 1-8 0.429,0.366 0.336 B 1-6 0.273, 0.277 10 A 1-10 0.482, 0.508 0.438 B 1-6 0.380,0.384 20 A 0.5-3 0.505, 0.522 0.464 B 1-4 0.404, 0.427 Pear  0 A Verylarge 0.049, 0.023 0.022 B Very large 0.013, 0.003  0.2 A 1-20 0.150,0.136 0.127 B 2-20 0.115, 0.110  0.5 A 1-8 0.187, 0.181 0.183 B 2-200.173, 0.194  1 A 1-8 0.240, 0.223 0.252 B 1-6 0.275, 0.270  5 A 1-20.577, 0.592 0.653 some larger B 1-3 0.776, 0.670 some larger *A and Bare duplicates.

TABLE 3 Shelf life stability Conc Emulsion Description (% w/v) BeforeAfter Centrifugation Gum arabic 0 Oil layer Oil layer Water layer Waterlayer 0.2 Oil film Oil layer Cream layer Water layer Water layer 0.5 Oilfilm Oil layer Cream layer Water layer Water layer 1 Cream layer Oillayer Water layer Cream layer Water layer 5 Cream layer Cream layerWater layer Water layer 10 Cream layer Cream layer Water layer Waterlayer 20 Cream layer Cream layer Water layer Water layer Pear 0 Oillayer Oil layer Water layer Water layer 0.2 Oil film Oil film Creamlayer Cream layer Water layer Water layer 0.5 Oil film Oil film Creamlayer Cream layer Water layer Water layer 1 Cream layer Cream layerWater layer Water layer 5 Cream Cream ¹Descriptions taken from duplicateemulsions.

TABLE 4 Composition and Selected Properties of Plant Cell Gums RELATIVEPROPORTIONS (%)¹ Caryophyllidae Dilleniidae Aizoacea ActinidiaceaeMalvaceae Cucurbitaceae Mesembryanthemum Actinidia deliciosa Hibiscusesculentus Sida rhombifolia Cucumis sativus COMPONENT “Pigface” Kiwifruit Okra Paddy's lucern Cucumber Protein & Ash² 26 41 30 52 33Xyloglucan 27 21 18 14 2 Galactoglucomannan — 6 — — — Glucomannan — — —3 3 3,6-Arabinogalactan (Type II) 18 21 17 23 15 Heteroxylan tr 2 1 2 54-Galactan (Type I) 1 — — — — Arabinan 4 1 1 2 31 Glucuronomannan — — —— 4 Rhamnogalacturonan/ 19 (55) 8 36 (32) 3 3 Galacturonan³Root-slime-like material — — — — — Emulsification 1.5 nt 56 10.6 13Droplet SIze 1% Gum⁴ Gelling 1% Gum⁵ NO NO NO NO NO Viscosity(Centipoise)⁶: 1/S 11 nt nt nt nt 100/S 7 1.9 2.5 1.7 1.6 RosidaeRoseace Rosa Fabaceae glauca Pyrus Mimosaceae Trigonella TrifoliumTrifolium Trifolium Rose communis Malus Acacia Acacia foenum- Medicagorepens pratense pratense Glycine Pauls Brown domesticus senegal senegalgraceum sativa White Red Red max COMPONENT Scarlet pear Apple (MS 5) (MS7) Fenugreek Alfalfa Clover Clover Clover Soy Protein + ash² 86 28 49 6041 58 56 48 76 Xyloglucan 3 17 17 36 52 9 7 4 26 5 5 Galactoglucomannan— — — — — — — — — — tr Glucomannan 1 — — — — — — 3 — — —3,6-Arabinogalactan 5 30 26 37 31 20 27 23 11 16 24 (Type II)Heteroxylan tr 2 2 3 4 5 2 2 1 1 1 4-Galactan (Type I) tr — — — — — — —— — — Arabinan tr 7 — — — 4 5 3 2 7 1 Glucuronomannan — 4 — — — — — — —11 — Rhamnogalacturonan/ 3 6 1 25 2 tr 9 (10) 5 (10) — 6 (7) trGalacturonan³ Root-slime-like material — — — — — — — — — — —Emulsification⁴ 52 27 9 nt nt 14 16 16 44.1 21.2 35 Droplet size (1%gum) Gelling at 1% gum⁵ NO NO nt nt nt NO NO NO NO NO NO Viscosity(Centipoise)⁶: 1/S nt 129 nt nt nt — nt nt nt nt nt 100/S 4.2 19 nt ntnt 2.1 1.3 1.6 1.7 1.7 1.9 Asteriade Solanaceae Nicotiana AsteraceaSolanum tuberosum Lycopersicon esculentum Nicotiana alataplumbaginifolia Cichorium intybus Letuca sativa COMPONENT Potato TomatoTobacco Ornamental Tobacco Chicory Lettuce Protein & Ash² 49 38 55 10 5270 Xyloglucan 15 9 8 32 5 4 Galactoglucomannan 12 10 7 13 — —Glucomannan — — — — 3 2 3,6-Arabinogalactan 14 16 13 10 20 6 (Type II)Heteroxylan 3 1 2 1 1 1 4-Galactan (Type I) — — — — 1 — Arabinan 2 1 2 —3 1 Glucuronomannan — — 1 22 (40) — — Rhamnogalacturonan/ 3 22 (11) 7 13(5)  14 (57) 13 Galacturonan³ Root-slime-like material — — — — — —Emulsification⁴ 35 32 56 15 13.6 7.4 Droplet size (1% gum) Gelling at 1%gum⁵ NO NO NO NO NO NO Viscosity (Centipoise)⁶: Rate 1s-1 9 8.7 nt 140nt nt Rate 100s-1 2.3 4.4 nt 55 1.7 1.9 Commelinidae (Monocot) PoaceaeLilidae (Monocot) Phleum Panicum Panicum Phalaris Oryza HordeumAgavaceae pratense millaceum millaceum aquaticus sativa vulgare ZeaPolianthus Polianthus Timothy Millet Millet Phalaris Rice Barley Maystuberosa tuberosa COMPONENT grass (MS3)⁷ (MS 9)⁷ (MS 6)⁷ Paddy Pelde⁷Schooner Maize (petal) (leaf) Protein & Ash² 12 39 35 58 70 66 34 54 51Xyloglucan 11 4 5 2 3 1 — 3 3 Galactoglucomannan — — — — — — — — —Glucomannan — 6 3 2 2 1 1 2 15 3,6-Arabinogalactan (Type II) 3 — — — — 47 6 20 Heteroxylan 11 7 14 9 4 16 44 2 6 4-Galactan (Type I) 1 7 12 trtr tr 3 1 — Arabinan — 3 6 1 2 5 — 1 2 Glucuronomannan — — — — — — — 25— Rhamnogalacturonan/ — — — — — tr — 2 1 Galacturonan³ Root-slime-likematerial 53 34 25 21 14 — — — — Emulsification: Droplet size (1% gum)⁴20 29 17 35 6.9 38.6 9.3 30 49 Gelling at 1% gum⁵ YES NO NO NO NO NO NONO NO Viscosity (Centipoise):⁶ 1/S 4500 1800 1800 10.4 nt nt nt 6.7 nt100/S 150 30 25 6.7 1.8 1.5 2 1.9 2.1 Table 4: FOOTNOTES ¹Calculated asthe sum of mol % of individual monosaccharide residues; tr means traceamounts; the percentage of components listed may add up to onlyapproximately 100% because the assumptions regarding structure may beinaccurate, such that the presence of minor functional groups such asmethyl or acetyl groups may not have been taken into account. ²Proteinand ash values in bold are assumed from total sugar determininations bycolorimetric analysis. ³Rhamnogalacturonan and galacturonan cannot beseparated by these analyses; the degree of methyl esterification inparentheses. ⁴Droplet size measured in μm; nt means not tested. ⁵Thepresence (YES) or absence (NO) of gelling is determined by visualobservation; nt means not tested ⁶Viscosity measured as a function ofshear rate in reciprocal seconds; nt means not tested ⁷Relativeproportion of polysaccharide deduced by comparison of linkage analysisfrom fractionated timothy grass biopolymer

TABLE 5 No. Age of No. sub- culture sub- cultures at cultures (suspen-harvest Species Explant Medium¹ (callus)² sion) (days) DicotsCaryophyllidae Aizoaceae Mesembryanthemum leaf MS2 25 6 13 DilleniidaeActinidiaccae Actinidia deliciosa fruit MS9   10+ 4 21 with seedMalvaceae Hibiscus esulentus seedling MS9  7 4 13 stem Sida rhomifoliaseedling MS9 20 4 14 stem Rosidae Rosaceae Pyrus communis ³ fruit pearBAL 24 7 11 Fabaceae Medicago sativa seed MS1 +   20+ 3  6 NOA Trifoliumrepens seed MS9   10+ 3 14 Trifolium pratense seed MS6  0 3 11 AsteridaeSolanaceae Solanum tuberosum growing MS6 18 5 18 points Lycopersiconfruit tomato   20+ 2 12 esculentum BAL Asteraceae Cichorium intybus DSM-LS   11+ 7 11 Gmbh⁴ Monocots Commelinidae Poaceae Phleum pratense seedMS +  0 3 14 2,4-D⁵ Panicum miliaceum seed MS3/MS9  2 4 14 LilidaeAgavaceae Polianthes tuberosa leaf MS17 10 3 18 ¹MS1 = MS powder with0.5 mg/L 2,4-D (2,4-dichlorophenoxyacetic acid); MS2 = MS powder with2.0 mg/L 2,4-D; MS3 = MS powder with 4.0 mg/L 2,4-D; MS6 = MS powderwith 4.0 mg/L 2,4-D and .075 mg/L mixed cytokinins; MS9 = MS powder with4.0 mg/L 2,4-D and 2.150 mg/L mixed cytokinins; MS17 = MS powder with2.0 mg/L NAA(1-naphthaleneacetic acid) and 2.0 mg/L BAP(6-benzylaminopurine); LS = LS powder with 0.2 mg/L 2,4-D and 0.04 mg/lNAA, see text Example 14; tomato BAL, see text Example 14; NOA =napthoxyacetic acid used at final concentration 0.5 mg/L; Concentrationof 2,4-D in Phleum pratense culture is 2.0 mg/L ²“+” means more, i.e.,10+ means more than 10. ³Brown pear. ⁴Commercial source of plant cellculture; Deutsche Sammlung von Mikroorganismen und Zelkulturen. ⁵Millethas been grown on both MS3 and MS9 with substantially the same results.The specific results shown were obtained on MS9.

TABLE 6 Polysaccharide Production in Selected Suspension Cell CulturesPOLYSAC- CHARIDE² CELL LINE EXPLANT MEDIUM¹ grams/litre DICOTSMesembryanthemum leaves MS2 3.1 sp “pigface ” Kiwi Fruit fruit MS1 0.06Actinidia deliciosa syn chinesis Okra seedling stem MS9 3.25 Hibiscusesculentis Paddy's lucern seedling stem MS9 0.5 Sida rhambifolia Pearfruit Pear BAL 3-4 Pyrus communis ³ Alfalfa seed MS + NOA 1.25 Medicagosativa White clover seeds MS9 0.85 Trifolium repens Red clover seeds MS61.9 Trifolium pratense Potato - red Solanum tuberosum growing points MS5and MS6 1.0 Tomato fruit Tomato BAL 1.75 Lycopersicon esculentumNicotiana exisiting CSV 3.5-4.5 Nicotiana culture⁴ plumbaginofoliaChicory DSM-Gmbh⁵ LS 1.6 Cichorium intybus Monocots Timothy Grass seedMS + 2,4-D 6.0 Phleum pratense Millet seeds MS9 Panicum miliaceumTuberose leaf MS17 2.7 Polyanthes tuberosa Tuberose petal TB1 1.9Polyanthes tuberosa ¹See footnote 1 Table 5 for media descriptions; CSVsee text Example 1.B; TB1 see text Example 14. ²The polysaccharideconcentration listed is the maximum concentration obtained in growthcycle measurements; Polysaccharide is typically harvested on the cultureday on which this maximum occurs; Tuberose petal gum was harvested onday 23. ³Brown pear ⁴The Nicotiana plumbaginifolia culture was obtainedfrom Paul Ebert of the School of Botany of the University of Melbourne(February 1989). ⁵Commercial source; Deutsche Sammlung vonMikroorganismem und Zelkulturen.

What is claimed is:
 1. A process for making a food product whichcomprises the step of adding a cultured plant cell gum of a plant of thefamily Aizoaceae to the food product.
 2. A process according to claim 1which comprises a step of adding a viscosifying agent, thickening agent,gelling agent, emulsifying agent, an emulsion stabilizing agent,suspending agent, encapsulating agent, enrobing agent, binding orcoating agent, or a texture modifier or any combination of thereof to afood product wherein the agent comprises a cultured plant cell gum of aplant of the family Aizoaceae.
 3. A process according to claim 1 whereinthe cultured plant cell gum is that of a plant of the genusMesembryanthemum.
 4. A process according to claim 1 wherein the culturedplant cell gum is that of the genera Mesembryanthemum or Carpobrutus. 5.A process according to claim 1 wherein an emulsifying agent is added tothe food product.
 6. The process according to claim 4 wherein the foodproduct is a soft drink.
 7. A food product that comprises a viscosifyingagent, thickening agent, gelling agent, emulsifying agent, an emulsionstabilizing agent, suspending agent, encapsulating agent, enrobingagent, binding or coating agent, or a texture modifier or anycombination of thereof wherein the agent comprises a cultured plant cellgum of the family Aizoaceae.
 8. A food product according to claim 7wherein the plant cell gum is that of a plant of the genusMesembryanthemum.
 9. A food product according to claim 7 wherein theplant cell gum is that of a plant of the genera Mesembryanthemum orCarpobrutus.
 10. A food product according to claim 7 comprising a plantcell gum that is an emulsifying agent or an emulsion stabilizing agent.11. A food product according to claim 7 which is a soft drink.
 12. Afood product according to claim 7 which comprises an encapsulated oilpowder.
 13. A food product according claim 7 which is a dietary fibersupplement.
 14. In an industrial, food, pharmaceutical or cosmeticmanufacturing process in which a plant exudate or plant extract gum isemployed as a thickening, emulsifying, suspending, waterproofing,gelling, protective colloid, stabilizing or coating agent, theimprovement wherein said plant exudate or extract gum is replaced with acultured plant cell gum of a plant of the family Aizoaceae.
 15. Aprocess according to claim 1 wherein the product is an edible capsuleparticularly an oil-containing capsule and wherein the plant cell gum isemployed to form the capsule.
 16. A process according to claim 15wherein the plant cell gum is that of a plant of the gunus Pyrus.
 17. Aprocess according to claim 1 wherein the cultured plant cell gum is thatof a plant of the genus Carpobrutus.
 18. A process according to claim 1wherein the cultured plant cell gum is that of a plant of the genusAptenia.
 19. A process according to claim 1 wherein the cultured plantcell gum is added to the food product to form a cloud emulsion.
 20. Afood product of claim 7 wherein the cultured plant cell gum is that of aplant of the genus Carpobrutus.
 21. A food product of claim 7 whereinthe cultured plant cell gum is that of a plant of the genus Aptenia. 22.A food product of claim 7 wherein the cultured plant cell gum is addedto form a cloud emulsion.